Pharmaceutical formulation containing DEMO for the treatment of cancer

ABSTRACT

Preparations comprising a capsule, tablet or other dosage form containing a core of different types of DFMO are provided. These preparations are capable of providing for the direct and constant delivery of DFMO to the entire GI tract or just the colon and rectum. The DFMO-containing granules include granules specially formulated to achieve rapid DFMO release, and granules formulated to achieve slower DFMO release and/or granules formulated for gastric, enteric or colorectal release. Methods of using the preparations to flood the GI tract with relatively constant levels of DFMO may thus be provided. The ratio of the (+) to the (−)-enantiomeric forms of DFMO in the granules will be controlled so as to enhance the pharmacological profile and reduce toxicity of the preparation relative to racemic DFMO. Preparations and methods for achieving systemic delivery as well as direct colon delivery of DFMO are also described.

[0001] This application is related to PCT No. US97/20424, filed Oct. 31,1997, designating the United States, the EP Application No. 97946915.2,filed Oct. 31, 1997, the Canadian Application No. 2/241,896, filed Oct.31, 1997, the Japanese Application No. 10-520855, filed Oct. 31, 1997,and the Mexican Application No. 985376, filed Jul. 1, 1998.

FIELD OF THE INVENTION

[0002] The present invention relates to a pharmaceutical formulation andits use for the treatment of cancer. More specifically, the presentinvention relates to alphadifluoromethylomithine (DFMO) containing oralpharmaceutical formulations having a varied release profile for thetreatment of cancer. These formulations may be modified for treatingspecific cancers.

BACKGROUND OF THE INVENTION

[0003] Both in vivo and in vitro, DFMO is an enzyme activatedirreversible inhibitor of ornithine decarboxylase (ODC) which isresponsible for the conversion of L-ornithine to putrescine, which inturn is converted to longer chain polyamines such as spermidine andspermine. These longer chain polyamines are required for cellularproliferation. Therefore, by inhibiting ODC, DFMO suppresses polyamineformation and consequently cellular proliferation. Aberrant andaccelerated cellular proliferation occurs in carcinogenic tissues. SinceDFMO is able to suppress polyamine formation, it is able to suppresscellular proliferation and ultimately to ameliorate or prevent cancer. Anumber of animal studies and/or human clinical trials relate to use ofracemic DFMO and specific neoplastic disorders. In addition, a clinicalstudy to determine the pharmacokinetics of racemic DFMO in healthy menhas been reported. (Haegele, 1981). Racemic DFMO was reported to have ashort elimination half-life, i.e., t_(½) is about 3.5 hours, as itundergoes rapid renal elimination. Peak plasma concentrations occurwithin about 6 hours after oral administration of racemic DFMOcontaining solutions. Mean total body clearance is about 1.20mL/min/LcKg, where mean renal clearance is about 0.99 mL/min/LcKgaccounting for 83% of drug elimination. The mean apparent volumedistribution is about 0.337 L/LcKg, corresponding to 24L for a 70 LcKgman. The amount of unchanged drug in 24-hour urine samples is about 44%after oral administration and about 80% after L.c. administration.

[0004] At a dose of about 3 g/m²V, a steady state level of DFMO,386-622, μM may be achieved. A DFMO dose of 2.25 g/m2 every six hourshas been recommended for Phase II studies in patients previously treatedwith cytotoxic drugs (Abeloff et al., 1984).

[0005] The maximally tolerated dose (MTD) of oral DFMO has also beenexamined (Abeloff et al., 1984). The MTD of a 4-day DFMO course givenorally, by CI, or by pulse IV infusions (Griffin et al., 1987) topatients with advanced solid tumors or lymphomas has also been studied.Some patients receiving twenty-four courses of oral DFMO on a 28-dayschedule developed thrombocytopenia (the DLT). Gastrointestinal sideeffects have also been observed in treated patients (Abeloff et al.,1984). Audiometric abnormalities is a further side effect associatedwith DFMO treatment (Griffin et al., 1987). No therapeutic responseswere noted in these patient populations.

[0006] A study by Griffin et al. (1987) compared routes (PO, CI and IV)and schedules (bolus and continuous infusions) of DFMO administration.Nausea and vomiting were the most frequent and severe toxicities noted,but this occurred mainly in patients receiving oral DFMO. Diarrhea wasalso observed in patients receiving oral DFMO. Mild leukopenia wasfurther observed with all routes of administration. Mildthrombocytopenia also occurred in some patients. No therapeuticresponses were reported with any route of drug administration.

[0007] The known minimum effective dose (MED) for racemic DFMO insignificantly reducing polyamine pools in vivo is about 0.43 g/day. Themaximum tolerated dose of these preparations reported is about 12g/m²/day (oral administration). The reported minimum toxic dose forthese racemic preparations of DFMO, in terms of ototoxicity, is about150 g/m² cumulative dose based upon 0.25-6.0 g/m²/day chronic oraladministration. GI toxicity occurs predominantly during P.O. rather thanI.V. administration of racemic DFMO preparations. (−)-DFMO has beenreported by some to be the enantiomer primarily responsible for ODCinhibition (Danzin, 1987). However, the side effects associated withDFMO have been traced to a particular enantiomeric form.

[0008] Tricalcium phosphate (TCP) and aluminum calcium phosphate (AlCAP)capsule formulations have been tested as implants in rats and proposedfor the treatment of trypanosomiasis. (Benghuzzi et al., 1988) A layeredtablet formulation comprising racemic DFMO and a slow release layercompressed to a rapid release layer has been tested for controllingfertility and gestation in rat and mouse models. (Bey et al., U.S. Pat.No. 4,309,442). Conventional release hard gelatin capsule and tabletformulations comprising racemic DFMO are also known and have been testedin rat, dog and/or mouse models for controlling gestation, treatingnon-malignant proliferative skin diseases and/or cancer chemoprevention.(Bey et al., U.S. Pat. No. 4,496,588).

[0009] A need continues to exist in the medical arts for formulationscapable of maintaining high plasma levels of DFMO during therapy inspite of its rapid clearance rate, without the toxic and/or non-pleasantside effects associated with available DFMO therapies.

SUMMARY OF THE INVENTION

[0010] It is an object of the present invention to provide an oral solidDFMO-containing pharmaceutical formulation for the treatment orprophylaxis of cancer. The formulations of the invention in some aspectsmaintain DFMO plasma levels in a patient below the minimum toxicconcentration or maximum tolerated concentration and above the minimumeffective concentration or minimum therapeutic concentration.

[0011] It is yet another object of the invention to provide a method oftreating or reducing the risk of cancer by administering to a patient aformulation as described above so as to provide to the gastrointestinal(GI) tract for an extended period of time a therapeutic amount of DFMO.

[0012] It is another object of the invention to provide aDFMO-containing dosage form having improved pharmacological activityrelative to racemic DFMO. The dosage form will comprise optically pure(−)-DFMO or (+)-DFMO or a defined ratio of (−)-DFMO: (+)-DFMO withreduced side effects or toxicity, enhanced therapeutic efficacy and/orimproved pharmacokinetics relative to racemic DFMO.

[0013] In one aspect, the invention provides an oral sustained releasepharmaceutical formulation for the treatment or prophylaxis ofcolorectal cancer comprising an oral dosage form. In some embodiments,this dosage form comprises a core and an outer layer surrounding thecore, where:

[0014] 1) the core comprises a rapid release granule and a slow releasegranule, and each granule comprises a therapeutic amount of (+)-DFMO,(−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO or pharmaceuticallyacceptable salts thereof; and

[0015] 2) the outer layer comprises a pH-responsive coating forcolorectal release of the core.

[0016] It is contemplated and within the scope of the invention that aformulation as described above will be useful for the inhibition ofcancer or tumor cell proliferation and the amelioration of colorectalcancer.

[0017] It is also contemplated and within the scope of the presentinvention that the pharmaceutical formulation may comprise racemic,optically pure or a defined ratio of the (+):(−) enantiomers of DFMO incombination with other therapeutic compounds for the treatment orprophylaxis of cancer.

[0018] It is also contemplated and within the scope of the presentinvention that the granules which comprise the dosage form core mayindividually or cooperatively exhibit zero-order, first-order orsecond-order DFMO release profiles in vivo or in vitro. The rapidrelease and slow release granules can also act cooperatively to providea patient being administered the oral sustained release formulation amean steady state plasma concentration level of (+)-DFMO, (−)-DFMO or adefined ratio of (+)-DFMO: (−)DFMO in the range of about 0.1 μM to about1000 μM.

[0019] It is contemplated and within the scope of the invention that thepresent formulation will provide a therapeutic benefit regardless of thelocation of the cancer.

[0020] In another aspect, the present invention provides an oralsustained release pharmaceutical formulation for the treatment orprophylaxis of colorectal cancer comprising an oral dosage form having acore and an outer layer surrounding the core, where:

[0021] 1) the core comprises a rapid release granule and a slow releasegranule, and each granule comprises a therapeutic amount of (+)-DFMO,(−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO or pharmaceuticallyacceptable salts thereof; and

[0022] a) the rapid release granule releases into the colorectal tract amajor portion of its DFMO within two hours after dissolution of theouter layer; and

[0023] b) the slow release granule releases into the colorectal tract amajor portion of its DFMO within eight hours after dissolution of theouter layer; and

[0024] 2) the outer layer comprises a pH-responsive coating or bacteriasensitive for colorectal release of the core at or above a pH of about6.

[0025] In another aspect, the present invention provides an oral solidmultiple drug release profile pharmaceutical formulation for thetreatment or prophylaxis of cancer. In some embodiments, these oralsolid multiple drug release profile pharmaceutical formulations comprisean oral dosage form having a core and an outer layer surrounding thecore for gastric release of the core, wherein the core comprisesDFMO-containing granules having different release characteristics; andthe granules comprise (+)-DFMO, (−)-DFMO or a defined ratio of (+)-DFMO:(−)-DFMO, or pharmaceutically acceptable salts thereof.

[0026] In another embodiment, the oral solid multiple drug releaseprofile pharmaceutical formulation for the treatment or prophylaxis ofcancer comprises: an oral dosage form having a core and an outer layersurrounding the core for gastric release of the core, where the corecomprises:

[0027] a gastric release granule;

[0028] an enteric release granule; and

[0029] a colorectal release granule; and

[0030] where each granule comprises a therapeutic amount of (+)DFMO,(−)-DFMO or a defined ratio thereof, or pharmaceutically acceptablesalts thereof.

[0031] It is contemplated and within the scope of the present inventionthat the dosage form may contain a variety of DFMO containing granuleshaving different release properties for delivery of DFMO throughout theGI tract. The granules maybe present in a range of different weightratios, may each contain a different amount of DFMO, and may themselvesbe comprised of other granules. The overall DFMO release profile of thismultiple drug release profile pharmaceutical formulation can approximatea zero-ordered controlled release profile.

[0032] It is intended that DFMO concentration in plasma will bemaintained at or above the minimum effective concentration for a majorportion of time during which DFMO-containing granules are present in apatient. The gastric release, enteric release and colorectal releasegranules can also act cooperatively to provide a patient beingadministered the oral solid multiple drug release profile pharmaceuticalformulation a mean steady state plasma concentration level of (+)-DFMO,(−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO in the range of about0.1μM to about 1000μM.

[0033] Following long-standing patent law convention, the terms “a” and“an” mean “one or more” when used in this specification.

[0034] Another aspect of the invention provides an oral sustainedrelease pharmaceutical formulation for the treatment or prophylaxis ofcolorectal cancer. In some embodiments, the formulation comprises anoral dosage form having a core and an outer layer surrounding the core,where:

[0035] 1) the core comprises a rapid release granule and a slow releasegranule, and each granule comprises a therapeutic amount of (+)-DFMO,(−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO or pharmaceuticallyacceptable salts thereof;

[0036] 2) the rapid release granule further comprises a binder;

[0037] 3) the slow release granule further comprises a polymer; and

[0038] 4) the outer layer comprises a pH-responsive coating forcolorectal release of the core at or above a pH of about 6.

[0039] In yet another aspect, the invention provides an oral solidmultiple drug release profile pharmaceutical formulation. In someembodiments, the formulation is employed for the treatment orprophylaxis of cancer. Some of these formulations may be defined ascomprising: an oral dosage form having a core and an outer layersurrounding the core for gastric release of the core, where the corecomprises:

[0040] a gastric release granule comprising a binder and an excipient;

[0041] an enteric release granule comprising a polymer suitable forenteric drug delivery; and

[0042] a colorectal release granule comprising a polymer which dissolvesat a pH greater than or equal to about 6;

[0043] wherein each granule comprises a therapeutic amount of (+)DFMO,(−)-DFMO or a defined ratio thereof, or pharmaceutically acceptablesalts thereof.

[0044] In yet another aspect, the present invention provides an oralsustained release pharmaceutical formulation for the treatment orprophylaxis of colorectal cancer in a patient. In some embodiments,these formulations comprise an oral dosage form having a core and anouter layer surrounding the core where:

[0045] the core comprises a rapid release granule and a slow releasegranule, and each granule comprises a therapeutic amount of (+)-DFMO,(−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO or pharmaceuticallyacceptable salts thereof; and

[0046] the outer layer comprises a pH-responsive coating for colorectalrelease of the core; wherein the slow release and rapid release granulesprovide a mean steady state plasma concentration level of total DFMOabove the minimum effective concentration for a major portion of thetime that slow release and/or rapid release granules containing(+)-DFMO, (−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO are presentin a patient administered the sustained release pharmaceuticalformulation.

[0047] In another aspect, the invention provides an oral solid multipledrug release profile pharmaceutical formulation for the treatment orprophylaxis of cancer. In some embodiments, the formulation may bedefined as comprising: an oral dosage form having a core and an outerlayer surrounding the core for gastric release of the core, where thecore comprises:

[0048] a gastric release granule;

[0049] an enteric release granule; and

[0050] a colorectal release granule;

[0051] wherein each granule comprises a therapeutic amount of (+)DFMO,(−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO, or pharmaceuticallyacceptable salts thereof, and the gastric release, enteric release andcolorectal release granules cooperatively provide a mean steady stateplasma concentration level of total DFMO above the minimum effectiveconcentration for a major portion of the time that gastric release,enteric release and/or colorectal release granules containing (+)-DFMO,(−)-DFMO or a defined ratio of (+)-DFMO: (−)-DFMO are present in apatient administered the multiple drug release profile pharmaceuticalformulation.

[0052] In yet another aspect, the present invention provides a sustainedrelease or multiple drug release profile pharmaceutical formulation asdescribed above that provides a mean steady state plasma concentrationlevel of total DFMO in the range of about 0.1 μM to about 1000 μM, orabout 1 μM to about 100 μM, or about 1 M to about 50 μM.

[0053] The components used in the present invention are available from avariety of commercial sources as follows:“EUDRAGIT™” polymers (RohmPharma, Germany), “AQUA-COAT™” and “AQUATERIC™” (F.M.C. Corp., PA),“SURELEASE”™ and “COATERIC™” (Colorcon, Inc., PA), “AQOAT™” (Shin-EtsuChemical Corp., Japan).

[0054] Unless otherwise indicated, all other chemicals were purchasedfrom Aldrich Chemicals (Milwaukee, Wis.) or Sigma Chemical Co. (St.Louis, Mo.). Racemic and enantiomeric forms of DFMO are available fromIlex Oncology.

[0055] As used herein, the term “granule” is taken to mean particle,crystal, minitablet, crystal, powder, particulate, or other similarsolid forms. The granules used in the invention may display diffusionand/or dissolution controlled release rate profiles according to thecomponents from and processes by which they are made.

[0056] As used herein, the term “adsorbent” is intended to mean an agentcapable of holding other molecules onto its surface by physical orchemical (chemisorption) means. Such compounds include, by way ofexample and without limitation, powdered and activated charcoal and thelike.

[0057] As used herein, the term “antioxidant” is intended to mean anagent which inhibits oxidation and thus is used to prevent thedeterioration of preparations by the oxidative process. Such compoundsinclude, by way of example and without limitation, ascorbic acid,ascorbyl palmitate, butylated hydroxyanisole, butylated hydroxytoluene,hypophophorous acid, monothioglycerol, propyl gallate, sodium ascorbate,sodium bisulfite, sodium formaldehyde sulfoxylate and sodiummetabisulfite and the like.

[0058] As used in the description of the present invention, a granule isdefined as, in some embodiments, an agglomerate, a pellet, a tablet, acollection of more than one particle, or a combination of these.

[0059] As used herein, the term “buffering agent” is intended to mean acompound used to resist change in pH upon dilution or addition of acidor alkali. Such compounds include, by way of example and withoutlimitation, potassium metaphosphate, potassium phosphate, monobasicsodium acetate and sodium citrate anhydrous and dihydrate and the like.

[0060] As used herein, the term “colorant” is intended to mean acompound used to impart color to liquid and solid (e.g., tablets andcapsules) pharmaceutical preparations. Such compounds include, by way ofexample and without limitation, FD&C Red No. 3, FD&C Red No. 20, FD&CYellow No. 6, FD&C Blue No. 2, D&C Green No. 5, D&C Orange No. 5, D&CRed No.8, caramel, and ferric oxide, red and the like.

[0061] As used herein, the term “flavorant” is intended to mean acompound used to impart a pleasant flavor and often odor to apharmaceutical preparation. In addition to the natural flavorants, manysynthetic flavorants are also used. Such compounds include, by way ofexample and without limitation, anise oil, cinnamon oil, cocoa, menthol,orange oil, peppermint oil and vanillin and the like.

[0062] As used herein, the term “sweetening agent” is intended to mean acompound used to impart sweetness to a preparation. Such compoundsinclude, by way of example and without limitation, aspartame, dextrose,glycerin, mannitol, saccharin sodium, sorbitol and sucrose and the like.

[0063] As used herein, the term “tablet antiadherents” is intended tomean agents which prevent the sticking of tablet formulation ingredientsto punches and dies in a tableting machine during production. Suchcompounds include, by way of example and without limitation, magnesiumstearate and talc and the like.

[0064] As used herein, the term “tablet binders” is intended to meansubstances used to cause adhesion of powder particles in tabletgranulations. Such compounds include, by way of example and withoutlimitation, acacia, alginic acid, carboxymethylcellulose sodium,compressible sugar (e.g., NuTab), ethylcellulose, gelatin, liquidglucose, methylcellulose, povidone and pregelatinized starch and thelike.

[0065] As used herein, the term “tablet and capsule diluent” is intendedto mean inert substances used as fillers to create the desired bulk,flow properties, and compression characteristics in the preparation oftablets and capsules. Such compounds include, by way of example andwithout limitation, dibasic calcium phosphate, kaolin, lactose,mannitol, microcrystalline cellulose, powdered cellulose, precipitatedcalcium carbonate, sorbitol, and starch and the like.

[0066] As used herein, the term “tablet direct compression excipient” isintended to mean a compound used in direct compression tabletformulations. Such compounds include, by way of example and withoutlimitation, dibasic calcium phosphate (e.g., DITAB) and the like.

[0067] As used herein, the term “tablet disintegrant” is intended tomean a compound used in solid dosage forms to promote the disruption ofthe solid mass into smaller particles which are more readily dispersedor dissolved. Such compounds include, by way of example and withoutlimitation, alginic acid, carboxymethylcellulose calcium,microcrystalline cellulose (e.g., AVICEL), polacrilin potassium (e.g.,AMBERLITE), sodium alginate, sodium starch glycollate, and starch andthe like.

[0068] As used herein, the term “tablet glidant” is intended to meanagents used in tablet and capsule formulations to reduce friction duringtablet compression. Such compounds include, by way of example andwithout limitation, colloidal silica, cornstarch, and talc and the like.

[0069] As used herein, the term “tablet lubricant” is intended to meansubstances used in tablet formulations to reduce friction during tabletcompression. Such compounds include, by way of example and withoutlimitation, calcium stearate, magnesium stearate, mineral oil, stearicacid, and zinc stearate and the like.

[0070] As used herein, the term “tablet capsule opaquant” is intended tomean a compound used to render a capsule or a tablet coating opaque. Maybe used alone or in combination with a colorant. Such compounds include,by way of example and without limitation, titanium dioxide and the like.

[0071] As used herein, the term “tablet polishing agent” is intended tomean a compound used to impart an attractive sheen to coated tablets.Such compounds include, by way of example and without limitation,carnauba wax, and white wax and the like.

[0072] It should be understood, that compounds used in the art ofpharmaceutical formulation generally serve a variety of functions orpurposes. Thus, if a compound named herein is mentioned only once or isused to define more than one term herein, its purpose or function shouldnot be construed as being limited solely to that named purpose(s) orfunction(s).

[0073] The term “unit dosage form” is used herein to mean a single ormultiple dose form containing a quantity of the therapeutic compoundcontaining formulation, said quantity being such that one or morepredetermined units are normally required for a single therapeuticdministration. In the case of multiple dose forms, such as scoredtablets, said predetermined unit will be one fraction, such as {fraction(1/2)}, of a scored tablet. “Halo” or “halogen” as used herein refers tofluoro, chloro, bromo, and iodo; and “counterion” is used to represent asmall, negatively charged species such as chloride, bromide, hydroxide,acetate, sulfate, and the like.

[0074] As used herein, the term DFMO is intended to meanalpha-difluoromethylornithine in its pharmaceutically acceptable saltand/or isomeric forms. (+)-DFMO is intended to mean a substantiallyoptically pure preparation of alpha-difluoromethylornithine having the(D)-configuration around the alpha-carbon of the molecule. (−)-DFMO isintended to mean a substantially optically pure preparation ofalpha-difluoromethylornithine having the (L)-configuration around thealpha carbon. By “a defined ratio of (+)-DFMO: (−)-DFMO” is meant aratio of the individual DFMO optical isomers in the range of about 5-95%wt: 95-5% wt., respectively. By “substantially optically purepreparation” is meant a preparation of a first enantiomer which containsabout 5% wt. or less of the opposite enantiomer. By “total DFMO” ismeant the total amount of DFMO present, i.e., the sum total of itsenantiomers, in plasma, for example.

[0075] Other features, advantages and embodiments of the invention willbe apparent to those skilled in the art from the following description,accompanying data and appended claims.

BRIEF DESCRIPTION OF THE DRAWINGS

[0076] The following drawings form part of the present specification andare included to further demonstrate certain aspects of the invention.The invention may be better understood by reference to one or more ofthese drawings in combination with the detailed description of specificembodiments presented herein.

[0077]FIG. 1. Representations of the approximate DFMO release profilesfor various embodiments of the multiple drug release profile formulation(compound preparation (a rapid release uncoated pellet of DFMO, onenteric coated rapid release pellet and colonic coated, rapid releasepellets in combination) in a solid dosage form is represented. Theplasma time release profile of DFMO after oral administration of theproduct containing the above compound is shown. The plasma time releaseprofile of DFMO after oral administration of the rapid gastric releasepreparation in a solid dosage form is demonstrated in FIG. 1. The rapidgastric release preparation is presented as solid squares (-▪-▪-▪-). Theplasma release profile of DFMO after oral administration of the entericcoated rapid enteric release preparation in a solid dosage form ispresented as solid circles (----). The colonic coated rapid releasepreparation in a solid dosage form is presented as open circles(-∘-∘-∘-). The “total” line is presented as a dashed line ( - - - ), andrepresents DFMO drug release in a composite tablet or capsule thatincludes the coated and uncoated particles in combination. Thecombination of these various pellets may be changed to provide thedesired drug delivery to target sites in the GI tract by modifying theamount of drug to be delivered with the desired coating to achieve thegastric, enteric or colorectal release of the agent.

[0078]FIG. 2. Representations of the approximate DFMO release profilesfor three embodiments of the sustained release formulation. The rapidcolorectal release profile is demonstrated as open circles (-∘-∘-∘-).The slow colonic release profile is demonstrated as open diamonds(-⋄-⋄-⋄-). The rapid release pellets in this profile (-⋄-⋄-⋄-) werefirst coated with EUDRAGIT™ 4110D RS 30D and RL 30D and then coated withEUDRAGIT™ 4110D. The composite total formulation provides a DFMO drugrelease profile that is presented in a graph as a dashed line ( - - - ).Modified ratios of these two types of coated granules may be created toprovide the desired drug delivery profile.

[0079]FIG. 3. The slow enteric release DFMO drug release labelledprofile comprises a slow release matrix pellet formulation as describedin Example 15, table 31. The slow enteric release coated preparationprovides a drug release profile as noted in the filled diamonds(-♦-♦-♦-). The rapid gastric release preparation is expected to providea DFMO drug release profile as noted in the solid squares (-▪-▪-▪-). Theslow colorectal release preparation will provide an expected DFMOrelease profile as presented in the open squares (-□-□-□-). The slowrelease matrix pellets were first coated with Opadry® II and thenEUDRAGIT™ 4110D. The in vitro disolution profile of these uncoated slowrelease matrix pellets appear at FIG. 18.

[0080]FIG. 4. Proposed in vivo plasma release of DFMO for matrix pelletscoated with EUDRAGIT™ 4110D is demonstrated on the graph as filleddiamonds (-♦-♦-♦-). The rapid release granules of DFMO were first coatedwith AQUACOAT® containing HPMC and then coated with EUDRAGIT™ 4110D(open circles, -∘-∘-∘-). The combination of these two pellets is alsoshown as the dashed line ( - - - ).

[0081]FIG. 5. Proposed in vivo plasma release profile of DFMO. Theproposed enteric coated rapid gastric release pellet is presented asfilled circles (----). The proposed slow release uncoated maxtixpellets provide a release profile presented as open triangles (-Δ-Δ-Δ-).The proposed colorectal coated slow release matrix pellets are presentedas open squares (-□-□-□-). The proposed DFMO drug release profile ispresented as a dashed line ( - - - ).

[0082]FIG. 6. Proposed in vivo plasma release profile of DFMO. Theproposed rapid gastric release coated pellet profile is presented asfilled squares (-▪-▪-▪-). The enteric coated slow release matrix pelletsare presented as solid diamonds (-♦-♦-♦-). The colorectal coated rapidrelease pellets are presented as open circles (-∘-∘-∘-).

[0083]FIG. 7. A first embodiment of a dosage form for the multiple drugrelease profile formulation. 1=hard gelatin capsule or caplet shapedtablet; 2=coated pellet; 3=matrix slow release pellet; 4=rapid releasepellet.

[0084]FIG. 8. A first embodiment of a dosage form for the sustainedrelease formulation. 5=coated pellet; 6=rapid release pellet;7=retardant coating or immediate release coating.

[0085]FIG. 9. Cross-sectional view of a matrix rapid release granule ofthe invention. 8=spherical pellet; 9=drug in pellet core.

[0086]FIG. 10. Cross-sectional view of a coated rapid release granule ofthe invention. 10=drug in core; 11=coating.

[0087]FIG. 11. Cross-sectional view of a multi-layered rapid releasegranule of the invention. 10=drug in core; 11=multi coated pellet;12=immediate release coating that may contain drugs; 13=coatings thatdissolve or disintegrate as a function of pH or microbial interaction.

[0088]FIG. 12. Cross-sectional view of a first tablet dosage form of thesustained release formulation embodiment of the invention. 15=drug inmatrix core for slow delivery to the GI tract; 16=drug in priming dosein HPMC or similar rapid release coating.

[0089]FIG. 13. Cross-sectional view of a second tablet dosage formcontaining sustained release coated pellets in a rapidly disintegratingtablet compact. 19=rapid release pellet; 18=sustained release coating(EUDRAGIT™ 4110 or EUDRAGIT™ 4110D); 17=tablet granulation containingDFMO for immediate release and absorption; 20=compressed tablet.

[0090]FIG. 14. Dissolution profile of uncoated rapid release pellets in500 ml, 0.1 N HCl using the USP method II.

[0091]FIG. 15. Dissolution profile in pH 1.0 and 6.8 of rapid releaseDFMO pellets coated with 8% EUDRAGIT™ L30D 55.

[0092]FIG. 16. Influence of pH on the dissolution properties of rapidrelease DFMO pellets coated with EUDRAGIT™ 4110D.

[0093]FIG. 17. Influence of pH on the release of DFMO from rapidrelease, DFMO core pellets coated first with 10% EUDRAGIT™ RS/RL30D(9:1) and then 8% EUDRAGIT™ 41100D as presented as open squares (

).

[0094]FIG. 18. Release profile in vitro of uncoated slow release matrixpellets at pH 7.4 (

) and pH of 1.0 (

). The data demonstrates that the release achieved with these pellets ispH independent.

[0095]FIG. 19. Release of DFMO from coated wax containing matrix pelletsat pH 1.0 and pH 6.8. (

)=EUDRAGIT™ 4110D coating; (

)=EUDRAGIT™ L30D-55). Proposed percent DFMO released from a preparationcomprising a core of a wax matrix pellet having a first coat of Opadry®II (as a subcoat), and a second coat of a EUDRAGIT™ 4110D or S100,illustrated with dashed, filled squares (

). Proposed percent DFMO released from a preparation comprising a coreof a wax matrix pellet with a first coat of Opadry® II (as a subcoat),and a second coat of EUDRAGIT™ L30D-55, is illustrated in the dashed,dotted line (

).

[0096] The subcoat of Opadry® II may comprise a 2% to 3% weight gain ofHPMC. The acrylic polymeric coating may then be applied by spraying thepolymer over the subcoat. The retardant polymer coat in some embodimentsmay be expressed as an amount that comprises about an 8% to an about 18%net weight gain. By way of example, these polymers may compriseEUDRAGIT™ L30D-55 or EUDRAGIT™ 4110D.

[0097] The release profiles in an acidic media reflect poor adhesion ofthe anionic polymers to the wax pellets containing DFMO. The rapidrelease of the drug in the absence of the subcoat was evident from bothpellet formulations in the acidic medium as demonstrated in FIG. 19.

[0098]FIG. 20. Demonstrates profile of a slow release matrix pelletformulation that was first coated with an ethyl cellulose dispersion(AQUACOAT®) containing HPMC, and then coated with EUDRAGIT™ 4110D.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0099] The present invention provides a variety of combinedvaried-release pharmaceutical formulations that include DFMO, andprovide for its delivery in preparations designed to be capable ofsustaining a desired release profile and/or plasma concentrationtailored to the pathology or condition being treated. The particularpreparations described below are examples of some of these formulationsand variations thereof in view of same are intended as within the scopeof the invention.

[0100] Dosage Form Outer Layer

[0101] The dosage form outer layer (7) which surrounds the dosage formcore comprising granules (5) and (6) will generally be insoluble ingastric juices and will release the core in a pH responsive fashion.Advantageously, the outer layer will release the core upon exposure to apH of about 6 or higher and will generally have released the dosage formcore contents directly into the colorectal tract of a patient. Bytargeting the colorectal tract for DFMO delivery, the presentformulation will find use in the treatment and/or prevention ofcolorectal cancer or other polyamine dependent colorectal disorders suchas premalignant polyps.

[0102] By “pH-responsive” is meant a pH-dependent fashion, i.e. theouter layer, when exposed to a certain pH, will deliver the dosage formcore to the colorectal tract. Advantageously, the outer layer will becomprised of a pH-responsive polymer that will dissolve when exposed toa pH greater than or equal to about 6; although, the pH-responsivepolymer may dissolve at a pH greater than or equal to about 5.

[0103] The outer layer will comprise, by way of example and withoutlimitation, a polymeric compound such as EUDRAGIT™ RS and EUDRAGIT™ RL.The EUDRAGIT™ products form latex dispersions of about 30D by weight.EUDRAGIT™ RS 30D is designed for slow release since it is not very waterpermeable as a coating and EUDRAGIT™ RS 30D is designed for rapidrelease since it is relatively water permeable as a coating. These twopolymers are generally used in combination. As contemplated herein, thepermissible ratios of EUDRAGIT™ RS 30D/EUDRAGIT™ RL 30D is about 10:0 toabout 8:2. Ethylcellulose or S100 or other equivalent polymers designedfor enteric or colorectal release can also be used in place of theEUDRAGIT™ RS/EUDRAGIT™ RL combination above. The outer layer can alsocomprise dyes, dissolution aids, colorants and pigments. Such componentsfor the outer layer are available from Colorcon (Delaware) or Crompton &Knowles (New Jersey).

[0104] Rapid Release Granule in the Dosage Form Core

[0105] With particular attention to FIG. 8, rapid release granules (6)will comprise part of the dosage form core which is surrounded in itsentirety by the outer layer (7). The rapid release granule willgenerally not be soluble in gastric juices and need not be soluble inenteric or colorectal fluids. It may release DFMO in either apH-dependent or pH-independent fashion and/or a zero-order, first-orderor second-order fashion. The rapid release granule will generally be adissolution controlled formulation. The rapid release granule mayprovide a patient with a loading dose of DFMO to quickly establish inthe patient a target DFMO plasma concentration level which is generallyabove the minimum effective concentration (MEC)of DFMO. Alternatively,the rapid release granule will permit the direct rather than systemicexposure of colorectal mucosa to DFMO.

[0106] The rapid release granule containing DFMO will advantageouslyhave released into the colorectal tract a major portion of its DFMOwithin two hours after dissolution of the dosage form outer layer.Although intended for colorectal release, it is possible this granulewill release some of its DFMO upstream in the GI tract if the dosageform outer layer has dissolved prematurely.

[0107] The rapid release granule will comprise DFMO and other compoundssuch as starch, talc, sugar, magnesium stearate, microcrystallinecellulose (MCC), lactose and pregelatinized starch. The rapid releasegranule can also comprise dissolution aids, stability modifiers,adsorption promoters, bioadhesive polymers and density modifiers.

[0108] The rapid release granule may be, by way of example and withoutlimitation, a quickly dissolving polymeric matrix (9) having DFMO (8)dispersed therein (FIG. 9), a DFMO core (10) surrounded by a rapidlydissolving coating (11) (FIG. 10), a multi-layered granule having anon-pareil core (12) surrounded by alternating layers of DFMO (13) and abinder (14) (FIG. 11) or a spheronized bead comprising DFMO, binder andexcipient integrally mixed therein.

[0109] Methods for the preparation of rapid release granules are wellknown in the art and depend upon whether the release is to bepH-dependent or pH-independent; or zero-order, first-order orsecond-order. One method for the preparation of these granules isdetailed in Example 2.

[0110] By “binder” is meant compounds such as hydroxypropylcellulose(HPC), hydroxypropylmethylcellulose (HPMC), methylcellulose and thelike. By “excipient” is meant an inactive adjuvant ingredient used inprocessing pharmaceutical solid dosage forms that aids in themanufacturing and processing or has a functional purpose afteradministration to a patient, e.g. disintegration aid. Binders include,by way of example and without limitation, compounds such asmicrocrystalline cellulose (MCC), carbohydrates, polysaccharides,pregelatinized starches, calcium phosphate, calcium sulfate, starchesand polymeric excipients.

[0111] Slow Release Granule in the Dosage Form Core

[0112] With particular attention to FIG. 8, slow release granules (5)will comprise part of the dosage form core which is surrounded in itsentirety by the outer layer (7). The slow release granule will generallynot be soluble in gastric juices and may release its DFMO in either apH-dependent or pH-independent fashion. The slow release granule canmaintain DFMO plasma concentration levels at or about the concentrationsachieved by the rapid release granule. By virtue of its release profile,the slow release granule will provide the colorectal mucosa with bothdirect and systemic exposure to DFMO.

[0113] The slow release granule containing DFMO will generally release amajor portion of its DFMO into the large intestine (colon) and therectum a major portion of its DFMO within eight hours after dissolutionof the dosage form outer layer. Although intended for colorectalrelease, it is possible this granule will release some of its DFMO priorto entering the colon if the dosage form outer layer has dissolvedprematurely.

[0114] The slow release granule will comprise DFMO and a polymer. Suchpolymers include EUDRAGIT™ S100, EUDRAGIT™ RS30D/EUDRAGIT™ RL30D,hydrophilic colloidal celluloses, aqueous dispersions of cellulose, suchas AQUA-COAT™ and SURELEASE, polysaccharides, such as sodium alginate,xanthan gum, proteins, gelatins, chitosan, clays, such asmerilonite,poly(ethylene oxide), bioadhesives, acrylic vinyl resins, such asCARBOPOL, polyacrylic acid resins, such as POLYCARBOPHYL, waxes,hydrogenated castor oil and high molecular weight vegetable oils.

[0115] The slow release granule may be, by way of example and withoutlimitation, a slow dissolving/releasing polymeric matrix having DFMOdispersed therein or a DFMO-containing core surrounded by a slowreleasing polymeric layer.

[0116] Methods for the preparation of slow release granules are wellknown in the art and depend upon whether the release is to bepH-dependent or pH-independent; diffusion or dissolution controlled; orzero-order, first-order or second-order. One method for the preparationof these granules is detailed in Example 3.

[0117] Dosage Formulation

[0118]FIG. 8 depicts one embodiment of the sustained release formulationof the present invention. In this caplet dosage form, the rapid releasegranules (6) together with the slow releases granules (5) are surroundedby a pH responsive outer layer (7). This embodiment comprises an outerlayer (7), a rapid release granule (6) and a slow release granule (5).This dosage form may be a capsule, gelcap or caplet. With particularattention to FIG. 12, the present dosage form may also be a tablet wherea core (15), comprising a slow releasing polymeric matrix having DFMOdispersed therein is surrounded by a layer (16), comprising DFMOintegrally mixed with a binder and an excipient, which is thensurrounded by a pH responsive outer layer (7). Referring now to FIG. 13,the dosage form can also be a tablet where the core (20) comprises DFMO(17) and slow release granules (5) dispersed therein. The slow releasegranules (5) comprise a pH responsive coating (18) surrounding a DFMOcore (19). The tablet core (20) is surrounded by a pH responsive coating(7).

[0119]FIG. 2 shows the expected plasma DFMO concentration profileexpected when one embodiment of the sustained release formulation of thepresent invention is used. MTC indicates the minimum toxic concentrationas determined by plasma concentration of DFMO. MEC indicates the minimumeffective concentration. It is intended that the mean plasma drugconcentration level, as indicated by the curved dashed line, will notdrop below the MEC for a major portion of the time in whichDFMO-containing granules remain in a patient. It is also intended thatthe two granules will act cooperatively to yield an approximate overallzero-order controlled release profile for the formulation as a whole.

[0120] The DFMO plasma concentration profile in a patient is the sumtotal of the release profiles of each given granule in a dosage form.Thus, various combinations of granules having different release profilescan act cooperatively to generate a DFMO plasma concentration profile asshown in FIG. 2.25.

[0121]FIG. 2 shows how a first rapid colorectal releasing granule and asecond slow colorectal releasing granule can act cooperatively to yieldthe desired overall DFMO plasma concentration profile for the sustainedrelease formulation of the invention. DFMO plasma concentration ismaintained between the MTC and MEC for a major portion of the time inwhich DFMO-containing granules are present in a patient. As per the areaunder the curve, the total amount of DFMO released by the rapid releasegranules can be less than that released by the slow release granules.

[0122]FIG. 4 shows how a first slow colorectal releasing granule and asecond rapid colorectal releasing granule can act cooperatively to yieldthe desired DFMO plasma concentration profile for the sustained releasedformulation of the invention.

[0123] It should be understood that specific release profiles for eachgranule will vary. It is contemplated and within the scope of theinvention that various release profile shapes may be attained by eachgranule without departing from the spirit and scope of the invention.

[0124] It should also be understood that the MEC will vary according tothe indication being treated, patient response, the DFMO form beingused, dosing regimen and a host of other reasons. It is generallyintended when referring to the MEC that the total amount of DFMO presentin plasma is being contemplated. When administered acutely, the MEC forDFMO will generally be higher than when it is administered chronically.When administered for preventing or reducing the risk of cancer, the MECfor DFMO will generally be lower than when it is administered fortreating or controlling the growth of cancer.

[0125] The MEC based upon mean steady state plasma concentration forDFMO will generally fall in the range of about 0.1 μM to about 1000 μMand preferably in the range of about 1 μM to about 100 μM and morepreferably in the range of about 1μM to about 50 μM. It should beunderstood that the maximum concentration level (Cmax) of DFMO willexceed the MEC but will generally be less than the MTC.

[0126] Thus according to one embodiment of the sustained releaseformulation of the invention, the slow release and rapid releasegranules will act cooperatively to provide a mean steady plasmaconcentration level for DFMO in the range of about 0.1 μM to about 1000μM and preferably in the range of about 1 μM to about 100 μM and morepreferably in the range of about 1 μM to about 50 μM.

[0127] The rapid and slow release granules of the invention may each bepresent in a wide range of amounts according to the formulation DFMOrelease profile desired. By way of example and without limitation, ifboth granules had the same amount of DFMO, it may be desirable toinclude the granules in the following ratio about 1:5 parts by weight(rapid: slow release granule) if predominantly direct flooding of thecolorectal tract by DFMO were desirable and if greater emphasis onsustaining a long DFMO release profile were desired. Conversely, theratio of about 5:1 parts by weight (rapid: slow release granule) wouldgive predominantly systemic exposure of the colorectal tract to DFMO andwould emphasize the rapid attainment of a maintenance plasma DFMOconcentration. The exact ratio chosen will depend upon individualpatient response to DFMO therapy, the extent of colorectal cancerprogression, the optical form of the DFMO administered, the actualdosage of DFMO in each granule or the desired dosing regimen.

[0128] Generally, each granule will be present in the range of about 1.0to about 9.0 parts by weight using about 10 parts by weight as the basisfor the total number of parts which the dosage form may contain, i.e.the granules will generally be present in the ratio of about 1.0-about9.0: about 1.0-about 9.0 parts by weight (rapid: slow release granule).Rather than to vary the relative amounts of rapid release and slowrelease it may be desirable to vary the amount of DFMO is each granule.

[0129] The dosage form of the present invention can be a tablet, caplet,gelcap or capsule, e.g. hard gelatin or soft gelatin capsule. When thesustained release formulation of the invention takes the form of agelcap, caplet or tablet, the granules may be held together by andcoated with a pH responsive layer that dissolves in colorectal fluidsthereby releasing the granules in the colorectal tract and directlyflooding it with DFMO.

[0130] When the dosage form is a capsule containing rapid and slowrelease particle, it may generally be prepared as follows. A capsuleouter layer comprised of an upper half and a lower half is preformed ofa material which is pH responsive and soluble in colorectal fluids. Thelower half is filled with core contents such as rapid and slow releasegranules and then capped (sealed) with the upper half to form a capsuleshaped dosage form. The relative sizes of the halves as well as thefinal size of the capsule outer layer generally depend upon the size andweight of the capsule core.

[0131] As used herein, the term “therapeutic compound” is taken to meana compound having the desired beneficial pharmacologic and therapeuticeffects in mammals. Advantageously, the therapeutic compound is alsocytoxic or cytostatic and indicated for the treatment or prophylaxis ofcancer.

[0132] The therapeutic compounds contemplated within the scope of theinvention may be in their free acid, free base, or pharmaceuticallyacceptable salt forms. They may be derivatives or prodrugs of a givencompound. -Loading of the therapeutic compounds into a pharmaceuticalformulation may be accomplished following well known techniques such asthose described [in] Remington's Pharmaceutical Sciences, 17th ed., MackPublishing Company, Easton, Pa., 1985, the disclosure of which is herebyincorporated by reference.

[0133] Therapeutic compound loading into the formulation may need to bevaried according to the pharmacological activity of the compound, theindication being treated, the targeted dosing regimen, the projectedmethod of administration, the integrity or stability of the finalformulation or other such reasons.

[0134] Multiple Drug Release Profile Formulation

[0135]FIG. 7 shows a capsule or tablet dosage form (1) for the multipledrug release profile formulation of the invention which comprises agastric release granule (4), an enteric release granule (3) and acolorectal coated pellet (2). The multiple drug release profileformulation will deliver DFMO throughout the entire gastrointestinal(GI) tract. This type of delivery will permit its use in treating,preventing and/or controlling the growth of a wide variety of cancersand tumors such as, by way of example and without limitation,neuroblastoma, colon carcinoma, leukemia, hepatoma, mammary sarcoma,small cell lung cancer, pancreatic tumor, Lewis lung carcinoma, B16murine melanoma, M3 murine adenocarcinoma, bladder carcinoma,endocervical carcinoma, epithelial cancer, chemically induced cancer,metastatic colorectal cancer, refractory childhood leukemia, cervicalintraepithelial neoplasia grade 3 (CIN III), hematological malignancies,acute and chronic myeloid leukemia, recurrent glioma and glioblastoma,solid tumor, lymphoma, mammary carcinoma, premalignant polyps andBarrett's esophagus.

[0136] Dosage Form Outer Layer

[0137] The dosage form outer layer which surrounds the dosage form corewill be soluble in gastric juices and may release the core in either apH-dependent or pH-independent fashion.

[0138] The dosage form outer layer will generally have released thedosage form core contents within two hours of administration of thedosage form to a patient.

[0139] The dosage form outer layer can be a film coating comprising apolymer such as, by way-of example and without limitation,hydroxypropylcellulose (HPC), ethylcellulose (EC),hydroxypropylmethylcellulose (HMPC), hydroxyethylcellulose (HEC), sodiumcarboxymethylcellulose(CMC), poly(vinyl pyrrolidone) (PVP),poly(ethylene glycol) (PEG), dimethylaminoethyl methacrylate—methacrylicacid ester copolymer, or ethylacrylate—methylmethacrylate copolymer(EA-MMA). The outer layer can also comprise dyes, dissolution aids,colorants, pigments, antiadhesives, surfactants, antifoaming agents andstabilizers.

[0140] Gastric Release Granule in Dosage Form Core

[0141] In one aspect, the invention provides a dosage form core thatcomprises a gastric release granule (4) and will be surrounded in itsentirety by the dosage form outer layer (1) (FIG. 7). The gastricrelease granule need not be soluble in gastric juices and may releaseDFMO in either a pH-dependent or pH-independent fashion.

[0142] The gastric release granule containing DFMO will generally havereleased into the stomach a major portion of its DFMO within two hoursof administration of the dosage form to a patient. Although intended forgastric release, it is possible this granule will release some of itsDFMO farther down the GI tract.

[0143] The gastric release granule will comprise DFMO and a binder suchas, by way of example and without limitation,hydroxypropylmethylcellulose (HPMC), ethylcellulose (EC),hydroxyethylcellulose (HEC), sodium carboxymethylcellulose (CMC),poly(vinyl pyrrolidone) (PVP), poly(ethylene glycol) (PEG),dimethylaminoethyl methacrylate—methacrylic acid ester copolymer, orethylacrylate—methylmethacrylate copolymer (GA-MMA) along with a fillerexcipient. The gastric release granule can also comprise dissolutionaids, stability modifiers, adsorption promoters, bioadhesive polymers,and plasticizers.

[0144] Methods for the preparation of gastric release granules are wellknown in the art and depend upon whether the release is to bepH-dependent or pH-independent; or zero order, first-order orsecond-order. One method for the preparation of the gastric releasegranule is detailed in Example 5.

[0145] Enteric Release Granule in the Dosage Form Core

[0146] In one aspect of the invention, the dosage form core willcomprise an enteric release granule (3) and will be surrounded in itsentirety by the dosage form outer layer (1) (FIG. 7). The entericrelease granule will generally not be soluble in gastric juices and mayrelease its DFMO in either a pH-dependent or pH-independent fashion.

[0147] The enteric release granule containing DFMO will generally havereleased into the small intestine a major portion of its DFMO withinabout six to eight hours of administration of the dosage form to apatient. Although intended for intestinal release, it is possible thisgranule will release some of its DFMO either prior to entering the smallintestine or farther down the GI tract.

[0148] The enteric release granule will comprise DFMO and a polymersuitable for enteric drug delivery such as, by way of example andwithout limitation, cellulose acetate phthalate (CAP), cellulose acetatetrimelletate (CAT), poly (vinyl acetate) phthalate (PVAP),hydroxypropylmethylcellulose phthalate (HP), poly(methacrylateethylacrylate) (1:1) copolymer (MA-EA), poly(methacrylatemethylmethacrylate) (1:1) copolymer (MA MMA), poly(methacrylatemethylmethacrylate) (1:2) copolymer, EUDRAGIT™ L 30D (MA-EA, 1:1),EUDRAGIT™ L100 55 (MA-EA, 3:1), hydroxypropylmethylcellulose acetatesuccinate (HPMCAS), SURETERIC (PVAP), AQUATERIC™ (CAP), shellac orAQOAT™ (HPMCAS). The enteric release granule can also comprisedissolution aids, stability modifiers, adsorption promoters, bioadhesivepolymers and plasticizers.

[0149] Methods for the preparation of enteric release granules are wellknown in the art and depend upon whether the release is to bepH-dependent or pH-independent; diffusion or dissolution controlled;zero-order, first-order or second-order; or rapid, slow or sustainedrelease. One method for the preparation of these granules is detailed inExample 6.

[0150] Colorectal Release Granule in the Dosage Form Core

[0151] In one aspect of the invention, the dosage form core willcomprise a colorectal release granule (2) and will be surrounded in itsentirety by the dosage form outer layer (1) (FIG. 7). The colorectalrelease granule need will generally not be soluble in gastric juices andmay release its DFMO in either a pH-dependent or pH-independent fashion.

[0152] The colorectal release granule containing DFMO will generallyhave released into the large intestine (colon) and the rectum a majorportion of its DFMO within about ten to sixteen hours of administrationof the dosage form to a patient. Although intended for colorectalrelease, it is possible this granule will release some of its DFMO priorto entering the colon.

[0153] The colorectal release granule will comprise DFMO and many of thesame or similar polymers employed in the enteric granule the maindifference being that a thicker coating of the polymer and that apolymer which dissolves at a pH greater than or equal to about 6 will beused. Targeted colonic delivery systems are known and employ materialssuch as hydroxypropylcellulose, microcrystalline cellulose (MCE, AVICEL™from FMC Corp.), poly (ethylene—vinyl acetate) (60:40) copolymer (EVACfrom Aldrich Chemical Co.), 2-hydroxyethylmethacrylate (HEMA), MMA,terpolymers of HEMA: MMA:MA synthesized in the presence of N,N′-bis(methacryloyloxyethyloxycarbonylamino)—azobenzene, azopolymers, entericcoated timed release system (TIME CLOCK® from Pharmaceutical Profiles,Ltd., UK) and calcium pectinate and the osmotic minipump system (ALZAcorp.). The colorectal release granule can also comprise dissolutionaids, stability modifiers, adsorption promoters, bioadhesive polymersand plasticizers.

[0154] Methods for the preparation of colorectal release granules arewell known in the art and depend upon whether the release is to bepH-dependent or pH-independent; diffusion or dissolution controlled;zero-order, first-order or second-order; or rapid, slow or sustainedrelease. One method for the preparation of these polymers is detailed inExample 7.

[0155] Sustained Release Granule

[0156] This granule is optionally present in the formulations of theinvention. When present, the sustained release granules will comprisepart of the dosage form core which is surrounded in its entirety by thedosage form outer layer. The sustained release granule may or may not besoluble in gastric or intestinal fluids and may release its DFMO ineither a pH-dependent or pH-independent fashion.

[0157] The sustained release granule containing DFMO can release DFMOthroughout the entire digestive tract or in targeted portions of the GItract.

[0158] The sustained release granule will comprise DFMO and, by way ofexample and without limitation, shellac, zein, ethylcellulose (EC),cellulose esters (such as acetate), silicone elastomers, acrylate estersor fats and waxes (such as beeswax, carnauba wax), hydrogenated castoroil and vegetable oils, cetyl alcohol, cetylstearyl alcohol, SURELEASE™(EC), AQUA-COAT™ (EC), EUDRAGIT™ NE 30D™ (EA-MMA), EUDRAGIT™ RL 30D™(poly [ethylacrylate—methylmethacrylate] triethylammonioethylmethacrylate chloride, EA-MMA-TEAE, 1:2:0.2) or EUDRAGIT™ RS 30D™(EA-MMA-TEAE, 1:2:0.1). The sustained release granule can also comprisedissolution aids, stability modifiers, adsorption promoters, bioadhesivepolymers and plasticizers.

[0159] Methods for the preparation of sustained release granules forrelease of therapeutic compound throughout the digestive tract are wellknown in the art and depend upon whether the release is to bepH-dependent or pH-independent; diffusion or dissolution controlled; orzero-order, first order or second-order release. One method for thepreparation of these granules is detailed in Example 8.

[0160] Multiple Drug Release Profile Capsule Dosage Form

[0161]FIG. 7 depicts one embodiment of this dosage form. Within thecapsule core are enteric (3), gastric (4), and colorectal (2) releasegranules. The outer layer (1) is shown as having a capsule shape but maypossess any desired shape.

[0162]FIG. 1 shows the expected plasma DFMO concentration profileexpected when one embodiment of the multiple drug release profileformulation of the present invention is used. It is intended that themean plasma drug concentration level will be maintained at or above theMEC for a major portion of the time in which DFMO-containing granulesremain in the GI tract. It is also intended that the granules will actcooperatively to yield overall an approximately zero-order controlledrelease profile. Three different types of rapid gastric, enteric andcolorectal release granules can combine to yield the desired overallDFMO plasma concentration profile.

[0163] Various combinations of rapid and slow gastric, enteric andcolorectal release granules can act cooperatively to yield the desiredoverall DFMO plasma concentration profile. FIG. 3 shows the combinationof a rapid gastric release granule, a slow enteric release granule and aslow colorectal release granule to yield the desired DFMO plasmaconcentration profile. FIG. 5 shows the combination of a slow gastricrelease granule, a rapid enteric release granule and a slow colorectalrelease granule to yield the desired overall DFMO plasma concentrationprofile. FIG. 6 shows the combination of a rapid gastric releasegranule, a slow enteric release granule and a rapid colorectal releasegranule to yield the desired overall DFMO plasma concentration profile.

[0164] Though specific release profiles are shown for each granule, itshould be understood that such profiles will vary. It is contemplatedand within the scope of the invention that other profile shapes may beattained by each granule without departing from the spirit and scope ofthe invention.

[0165] According to one embodiment of the multiple drug release profileformulation of the invention, the gastric release, enteric release andcolorectal release granules will act cooperatively to provide a meansteady state plasma concentration level of total DFMO in the range ofabout 0.1μM to about 1000μM and preferably in the range of about 1 μM to100μM and more preferably in the range of about 1 μM to about 50 μM.

[0166] The gastric, enteric and colorectal release granules of theinvention may each be present in a wide range of amounts according tothe overall DFMO release profile desired. By way of example and withoutlimitation, it may be desirable to include the granules in the followingratio 1:1:5 parts by weight (gastric: enteric: colorectal releasegranule) if predominantly direct flooding of the patient by DFMO weredesirable, or in the ratio 5:3:2 parts by weight (gastric: enteric:colorectal release granule) if predominantly systemic exposure of thepatient to DFMO were more desirable. The exact ratio chosen will dependupon individual patient response to DFMO therapy, toxicity, the extentof cancer progression, the optical form of the DFMO administered, theactual dosage of DFMO in each granule, the desired dosing regimen or thetype of cancer being treated.

[0167] Generally, each granule will be present in the range of about 1.0to about 9.0 parts by weight using 10 parts by weight as the basis forthe total number of parts by weight which the formulation may contain,i.e. the granules will generally be present in the ratio of about1.0-9.0: about 1.0-9.0: about 1.0-9.0 parts by weight (gastric: enteric:colorectal release granule).

[0168] When the multiple drug release profile formulation of theinvention takes the form of a capsule the gastric, enteric, sustainedand colorectal release granules will be found in their entirety enclosedwithin an outer layer, e.g. hard gelatin capsule, formulated for rapidrelease.

[0169] Methods for the preparation of the dosage forms just mentionedare well known in the art and depend upon whether release of the dosageform core is to be pH-dependent or pH independent, the type of granuleswithin the dosage form, and upon the desire to release the dosage formcore contents into the stomach, intestines or colon. Generally and asadvantageously employed herein, the dosage form has an outer layersurrounding a core which contents are released into the stomach. Amethod for preparing such a dosage form generally proceeds as follows. Acapsule outer layer comprised of an upper half and a lower half ispreformed of a material which is soluble in gastric fluids. The lowerhalf is filled with capsule core contents such as gastric, enteric andcolorectal release granules and then capped (sealed) with the upper halfto form a capsule dosage form. The relative sizes of the halves as wellas the final size of the capsule outer layer generally depend upon thesize and weight of the capsule core. One method for the preparation ofthis formulation is detailed in Example 10.

[0170] When the multiple drug release profile formulation of theinvention takes the form of a gelcap, caplet or tablet, the gastric,enteric and colorectal release granules may be held together by andcoated with an outer layer that dissolves in gastric fluids therebyreleasing the granules in the stomach. The outer layer can be a hardgelatin capsule comprised of gelatin, glycerin and sorbitol or othersuitable plasticizers.

[0171] Pharmaceutical Formulation and Administration

[0172] The pharmaceutical formulation of the present invention isintended for oral administration and may be provided in a variety ofways. Any ingredients used in the present formulation should not degradeor decompose a significant portion of the DFMO or other therapeuticcompound(s) used prior to administration.

[0173] The solid unit dosage form of the invention will comprise DFMOand can be combined with conventional carriers, for example, binders,such as acacia, corn starch or gelatin; disintegrating agents, such as,corn starch, guar gum, potato starch or alginic acid; lubricants, suchas, stearic acid or magnesium stearate; and inert fillers, such aslactose, sucrose or corn starch.

[0174] The dosage form may also comprise adsorbents, antioxidants,buffering agents, colorants, flavorants, sweetening agents, tabletantiadherents, tablet binders, tablet and capsule diluents, tabletdirect compression excipients, tablet disintegrants, tablet glidants,tablet lubricants, tablet or capsule opaquants and/or tablet polishingagents.

[0175] For gelcap preparations, the pharmaceutical formulation mayinclude oils, for example, fixed oils, such as peanut oil, sesame oil,cottonseed oil, corn oil and olive oil; fatty acids, such as oleic acid,stearic acid and isostearic acid; and fatty acid esters, such as ethyloleate, isopropyl myristate, fatty acid glycerides and acetylated fattyacid glycerides; with alcohols, such as ethanol, isopropanol, hexadecylalcohol, glycerol and propylene glycol; with glycerol ketals, such as2,2-dimethyl-1,3-dioxolane-4 methanol; with ethers, such aspoly(ethylene glycol) 450, with petroleum hydrocarbons, such as mineraloil and petrolatum; with water, or with mixtures thereof; with orwithout the addition of a pharmaceutically suitable surfactant,suspending agent or emulsifying agent.

[0176] Oils can also be employed in the preparation of formulations ofthe soft gelatin type. Water, saline, aqueous dextrose and related sugarsolutions, and glycerols may be employed in the preparation ofsuspension formulations which may suitably contain suspending agents,such as pectin, carbomers, methyl cellulose, hydroxypropyl cellulose orcarboxymethyl cellulose, as well as buffers and preservatives. Soaps andsynthetic detergents may be employed as surfactants and as vehicles fordetergent compositions. Suitable soaps include fatty acid alkali metal,ammonium, and triethanolamine salts. Suitable detergents includecationic detergents, for example, dimethyl dialkyl ammonium halides,alkyl pyridinium halides, and alkylamine acetates; anionic detergents,for example, alkyl, aryl and olefin sulfonates, alkyl, olefin, ether andmonoglyceride sulfates, and sulfosuccinates; nonionic detergents, forexample, fatty amine oxides, fatty acid alkanolamides, andpoly(oxyethylene) -block poly (oxypropylene) copolymers; and amphotericdetergents, for example, alkyl β-aminopropionates and 2-alkylimidazolinequaternary ammonium salts; and mixtures thereof.

[0177] Various other components, not listed above, may be added to thepresent formulation for optimization of a desired DFMO release profileincluding, by way of example and without limitation,glycerylmonostearate, nylon, cellulose acetate butyrate, d, 1-poly(lactic acid), 1,6-hexanediamine, diethylenetriamine, starches,derivatized starches, acetylated. monoglycerides, gelatin coacervates,poly (styrene—maleic acid) copolymer, glycowax, castor wax, stearylalcohol, glycerol palmitostearate, poly (ethylene), poly (vinylacetate), poly (vinyl chloride), 1,3-butylene-glycoldimethacrylate,ethyleneglycol dimethacrylate and methacrylate hydrogels.

[0178] Since the present formulation may comprise a variety of granules,it is contemplated that a combination of rapid acting, short-acting,fast-releasing, long-acting, gastric release, enteric release,colorectal release, sustained release, controlled release or slowrelease granules may be used in the present invention.

[0179] The course and duration of administration of and the dosagerequirements for the formulation of the present invention will varyaccording to the subject being treated, the formulation used, the methodof administration used, the severity and type of cancer being treated,the coadministration of other drugs and other factors.

[0180] Although each unit dosage form (capsule, tablet, gelcap orcaplet) contains therapeutically effective amounts of DFMO, it may benecessary to administer more than one such unit dosage form in order toobtain the full therapeutic benefit of the DFMO. More particularly,since DFMO may require moderately high doses, vide supra, for preventingand treating colorectal cancer, it is very likely that more than onecapsule, tablet, caplet or gelcap will need to be administered to apatient in order to obtain the full therapeutic benefit of DFMO.

[0181] For example, consider that the average 70 Kg man has a bodysurface area of 1.73 m². If DFMO is administered at a dosage of up toabout 3 g/m²/day, then a patient would have to receive about 5 g ofDFMO/day, about 10 tablets containing 0.5 g of DFMO. Correspondingly, ifthe dosage administered is about 0.25 g/m²/day, then a patient wouldhave to receive about 0.4 g/day, about 1 tablet containing 0.5 g ofDFMO.

[0182] It is intended that the formulation of the invention willmaintain DFMO plasma levels within the limits of MTC and MEC for a majorportion of the time during which dosage form granules are present in apatient. By the term “major portion” is meant at least about 50%.

[0183] The therapeutic compound contained within the formulation may beformulated as their pharmaceutically acceptable salts. As used herein,“pharmaceutically acceptable salts” refer to derivatives of thedisclosed compounds wherein the parent pharmacologically active compoundis modified by making acid or base salts thereof. Examples ofpharmaceutically acceptable salts include, but are not limited to,mineral or organic acid salts of basic residues such as amines; alkalior organic salts of acidic residues such as carboxylic acids; and thelike. The pharmaceutically acceptable salts include the conventionalnon-toxic salts or the quaternary ammonium salts of the parent compoundformed, for example, from non-toxic inorganic or organic acids. Forexample, such conventional non-toxic salts include those derived frominorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfonic,sulfamic, phosphoric, nitric and the like; and the salts prepared fromorganic acids such as amino acids, acetic, propionic, succinic,glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic,maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic,sulfanilic, 2-acetoxybenzoic, fumaric, toluenesulfonic, methanesulfonic,ethane disulfonic, oxalic, isethionic, and the like.

[0184] The pharmaceutically acceptable salts of the present inventioncan be synthesized from the parent pharmacologically active compoundwhich contains a basic or acidic moiety by conventional chemicalmethods. Generally, such salts can be prepared by reacting the free acidor base forms of these compounds with a predetermined amount of theappropriate base or acid in water or in an organic solvent, or in amixture of the two. Generally, nonaqueous media are preferred. Lists ofsuitable salts are found in Remington's Pharmaceutical Sciences, 17thed., Mack Publishing Company, Easton, Pa., 1985, p. 1418, the disclosureof which is hereby incorporated by reference.

[0185] The phrase “pharmaceutically acceptable” is employed herein torefer to those compounds, materials, compositions, and/or dosage formswhich are, within the scope of sound medical judgment, suitable for usein contact with the tissues of human beings and animals withoutexcessive toxicity, irritation, allergic response, or other problem orcomplication, commensurate with a reasonable benefit/risk ratio.

[0186] The compounds herein described may have asymmetric centers. Allchiral, diastereomeric, and racemic forms are included in the presentinvention. Many geometric isomers of olefins, C═N double bonds, and thelike can also be present in the compounds described herein, and all suchstable isomers are contemplated in the present invention unless thespecific stereochemistry or isomer form is specifically indicated. Itwill be appreciated that certain compounds of the present inventioncontain an asymmetrically substituted carbon atom, and may be isolatedin optically active or racemic forms. It is well known in the art how toprepare optically active forms, such as by resolution of racemic formsor by synthesis, from optically active starting materials. Also, it isrealized that cis and trans geometric isomers of the compounds of thepresent invention are described and may be isolated as a mixture ofisomers or as separated isomeric forms.

[0187] The racemic and optically pure forms of DFMO may be preparedaccording to the methods described by Bey et al. (U.S. Pat. No.4,413,141), Sjoerdsmannetal. (U.S. Pat. No. 4,399,151), Bey et al. (U.S.Pat. No. 4,438,270), ibid (U.S. Pat. No. 4,560,795), ibid (10 U.S. Pat.No. 4,743,691), ibid (U.S. Pat. No. 4,866,206), Au et al. (EP 357029AZ), Wagner et al. (Anal. Biochem. (1987) 164(1), 102-16), Lindner etal. (J. Liq. Chromatogr. (1986) 9 (2-3), 551-71) and Aldous et al. (J.Chromatogr. (1986) 357(2) 335-9) which references are herebyincorporated in their entirety herein.

[0188] The mean steady state plasma concentration level of racemic DFMOcan be determined according to the methods described in Smithers (Pharm.Res. (1988) 5, 684-686), Bitonti et al. (Biochem. Pharmacol. (1986), 35,351-354) and Grove et al. (J. Chromatogr. (1981), 223, 409-416) thedisclosures of which are hereby incorporated in their entirety herein.

[0189] The mean steady state plasma concentration level of theindividual DFMO enantiomers can be determined according to the methodsdescribed in Schmitt-Hoffinann (Annual Report of the CIFRE Convention,1987) the disclosure of which is hereby incorporated in its entiretyherein.

EXAMPLE 1 Determination of Racemic DFMO in Plasma

[0190] An internal standard (IS), 4-amino-3-hydroxybutyric acid (SeeFIG. 1 for structure of DFMO and IS) is added to 100 ul of plasmasample. The plasma is deproteinized by the addition of methanol. Aftercentrifugation the supernatant is transferred to a WISP® vial (0.9 mL,designed for the 96 position carousel). Phosphate buffer of pH =7.5 isadded to it. DFMO standards in plasma are prepared similarly. Thesamples and standards are analyzed by pre-column derivatization with9-phthalaldehyde (OPA). This is accomplished by Water's Auto-Tagtechnique which uses the WISP® to alternately inject reagent and sampleunder zero flow conditions. A controller activates the solvent pumps andthe reaction mixture is chromatographed by gradient elution. Detectionis by fluorescence and data acquisition and calculations are by CALS. Alinear curve regression of the peak area ratios of DFMO to IS VS. DFMOconcentrations is used to assign concentrations to unknown samples. Thisprocedure is used for the determination of DFMO in plasma over theconcentration range of 0.5 to 80 ug/mL. Complete details of the methodare described in the appendix. Materials and Reagents Ethanol -absolute, 200 proof, 9A Methanol (MeOH) - Burdick & Jackson, glassdistilled (B&H) Isopropanol (IPA) - (B&J), glass distilled acetonitrile(ACN) - (B&J), glass distilled 0.1 M phosphate Buffer, pH = 7.5 (A) .1 Msodium hydrogen phosphate (Na2HPO4) (B) .1 M sodium dihydrogen phosphate(NaH2PO4) (A) and (B) are combined in a ratio of approximately 13 to 3to give a pH of 7.5

[0191] O-Phthalaldehyde Reagent (OPA)-10.0 mg of o-phthalaldehyde (SigmaChemical Co.) is dissolved in 1 mL of ethanol. One hundred mcL of2-mercaptoethanol (Sigma Chemical Co.) and 10 mls of 0.1M phosphatebuffer, pH=7.5, is added to the ethanol solution.

[0192] Standard Solutions: Eflornithine. MDL 71 782A

[0193] Stock Solution

[0194] Accurately weight 50 mg of compound into a 25 mL volumetric flaskand 1.s. with glass distilled water (2 mg/mL).

[0195] Working Solutions A, B, C

[0196] A. Pipette 5 ml of stock solution into a 100 ml volumetric flaskand 1.s. with plasma (100 ug/ml).

[0197] B. Pipette 1 ml of stock solution into a 100 ml volumetric flaskand q.s. with plasma (20 ug/ml).

[0198] C. Pipette 5 ml of working solution B (above) into a 50 mlvolumetric and q.s. with plasma (2 ug/ml).

[0199] Internal Standard Solution:

[0200] 4-Amino-3-hydroxybutyric acid (Aldrich Chemical Co.) Weight 25 mginto a 100 ml volumetric flash and q.s. with glass distilled water.

[0201] Equipment

[0202] Centrifuge—Beckman Model TJ-6, Beckman Instrument Co.

[0203] Vortex—Genie Mixer, Scientific Instruments Inc.

[0204] Instrumentation

[0205] Analysis are performed on a Waters HPLC System(Millipore, Waters'Chromatography Division) consisting of a Model 720 System Controller,two Model 510 pumps and a WISP\, Model 710B auto-injector. Thechromatography column is a Waters' C18, 5 micron, Radial-Pak cartridge,preceded by a pre-column (Upchurch Scientific, Inc.) packed with WatersBondapak C18/corasil, 37-50 micron particle size. The fluorometer is aKratos Model FS970 (Schoeffel Instrument Division) operated at anexcitation of 335 nm with a 418 nm cut-off filter.

[0206] Procedure

[0207] I. Preparation of Standard Curve

[0208] 1. A series of nine standards, ranging in concentration from 80to 0.5 mcg/mL, and a blank are prepared as follows: TABLE 1 Volume (mL)of Working Std. Sample Solution Volume (mL) of Conc. No. A, B or C BlankPlasma ug/mL 1 4 (A) 1 80 2 3 (A) 2 60 3 2 (A) 3 40 4 5 (B) 0 20 5 3 (B)3 10 6 2 (B) 6 5 7 5 (C) 0 2 8 3 (C) 3 1 9 2 (C) 6 0.5 10 0     5 0

[0209] 2. One hundred mL aliquots of each standard are assayed induplicate. The aliquot is placed in a 13×100 mm test tube.

[0210] II. One hundred mcL of sample is placed in a 13×100 mm test tube.The sample is assayed in duplicate.

[0211] III. Analysis of Standard and Samples

[0212] A. Sample Preparation

[0213] 1. Using a 1.0 mL Hamilton syringe with a PB60 repeatingdispenser, add 20 uL (56 ug) of the IS to each standard and sample.

[0214] 2. Add 400 uL of methanol and mix on Vortex mixer to insurecomplete precipitation of proteins.

[0215] 3. Centrifuge of 30 minutes at approximately 2,000 rpm.

[0216] 4. Remove the supernatant to a 0.9 mL WISP® vial designed for the96 position carousel. Add 200 uL of 0.02 M phosphate buffer, pH=7.5.

[0217] 5. Fill a 0.9 mL WISP® vial with OPA reagent, cap and place inthe No. 1 position in the carousel.

[0218] 6. Under zero flow conditions 20 ul of OPA reagent is injectedfollowed by 15 uL of sample.

[0219] B. HPLC Operations and Conditions

[0220] Column: Radial-Pak, C18, 5 micron Nova)

[0221] Pre-column: 2 mm×2 cm, C18/Corasil, 37-50 microns.

[0222] Fluorometer: 335 nm excitation

[0223] 418 nm cut-off filter

[0224] Mobile Phases:

[0225] A. 92% 0.1 M Phosphate, pH=7.5,5% MeOH, 3% IPA

[0226] B. 80% MeOH, 10% H₂O, 5% ACN, 5% IPA

[0227] Pump Controller Program: TABLE 2 Time (min) Flow Rate (ml/min) %A % B Curve Init 0 80 20 11  2 0.2 80 20 11  3 1.5 80 20 11  5 1.5 80 2011 20 1.5 80 50 06 25 1.5 80 50 11 26 1.5 80 20 06 33 0 80 20 11

[0228] (There is an initial 3 min isocratic flow of 80% A and & 20% B.The flow rate the first minute is 0.2 ml/min. It changes to 1.5 ml/minand remains as such for the rest of the program. The linear gradient isto 50%A, 50%B over 15 min, after which the mobile phase changes to theinitial conditions and the column equilibrates for seven minutes beforethe next injection.)

[0229] WISP®

[0230] The WISP® is programmed to inject the OPA reagent, in positionno. 1 in the carousel, prior to each sample injection. This isaccomplished through programming system messages 82 and 72 as 8201 and7201 (WISP® Operator's Manual, p.4.4, section 4.3.1). The programmedinjection times for the OPA reagent and sample are 1 min and 34 minrespectively. The sample loop has a 2.0 ml capacity which acts as amixing chamber for sample and reagent.

EXAMPLE 2 Preparation of DFMO-Containing Rapid Release Granule

[0231] (−)-DFMO (100 g) and microcrystalline cellulose (MCC, AVICELPH101, 100 g) are mixed thoroughly. A sufficient amount of water to makea wet mass is added to the mixture which is subsequently extruded andspheronized according to well known procedures in the art. The pelletsare screened (size 14 to 20 mesh) and dried at 40 C. for 24 hours.Poly(vinyl pyrrolidone) (PVP, 2% by wt of total mass) can optionally beincluded in the formulation. Increasing PVP will generally lengthen therelease profile of the formulation.

EXAMPLE 3 Preparation of DFMO-Containing Slow Release Granule

[0232] The granules are prepared similar to Example 9. Thus, DFMO (500g), MCC (500 g) and EUDRAGIT™ RS 30D (35-50 g) are mixed. To thismixture is added sufficient water to yield a 30% wt. suspension. To thesuspension is added TEC (10% wt. based on dry polymer weight ofEUDRAGIT™) to yield a dispersion which is wet granulated and dried toremove as much water as possible. The particles are then ground into afine powder.

EXAMPLE 4 Preparation of DFMO-Containing Capsule Comprising Rapid andSlow Release Granules

[0233] The following procedure details the preparation of the dosageform described by FIG. 2. Rapid release granules (500 g preparedaccording to Example 2) and slow release granules (750 g preparedaccording to Example 3) are thoroughly mixed. The mixture is used tofill 2000 hard gelatin capsules according to procedures well known inthe art.

EXAMPLE 5 Preparation of DFMO-Containing Gastric Release Granule

[0234] The same method detailed in Example 2 can be employed here toprepare a rapid gastric release granule. Alternatively, a slow gastricrelease granule can be prepared as follows. DFMO (600 g), MCC (350 g)and HPC (50 g) are mixed thoroughly. To the mixture is added sufficientwater to make a wet mass which is extruded and then spheronized usingprocedures well known in the art. The particles are then dried andground.

EXAMPLE 6 Preparation of DFMO-Containing Enteric Release Granule

[0235] Rapid release.

[0236] A latex dispersion is prepared as follows. To EUDRAGIT™ L 30D-55(1000 g, 15% wt in water) is added a plasticizer (15% wt of dry polymerweight in the EUDRAGIT™) while mixing for 1 to 24 hours. Plasticizerssuch as triethylcitrate, tributylcitrate, acetyltributylcitrate ordibutylsebacate can be used. To this mixture is added talc (50% wt ofdry polymer in the EUDRAGIT™) or glycerylmonostearate (10% wt of drypolymer in the EUDRAGIT™) to form a dispersion. The rapid releasegranules prepared in Example 2 are coated in a fluidized bed with thelatex dispersion until a 10-15% wt increase in granule weight isachieved. The fluidized bed inlet air temperature is adjusted to about40-45 C. and the outlet air temperature is adjusted to about 30-35 C.with a spray rate of about 2 g/min.

[0237] Slow Release.

[0238] Granules prepared according to Example 3 are coated withEUDRAGIT™ L 30D (10-12% wt.) or AQUATERIC (CAP, 10% wt., plasticizedwith TEC) until a 25-30% wt. increase in granule weight is achieved.

EXAMPLE 8 Preparation of DFMO-Containing Colorectal Release Granule

[0239] Rapid Release.

[0240] A dispersion is prepared as follows. To EUDRAGIT™ S 100 (1000 g,10% wt in water) is added a plasticizer (10% wt of dry polymer weight inthe EUDRAGIT™ ) while mixing for 1 to 24 hrs. Plasticizers such astriethylcitrate, tributylcitrate, acetyltributylcitrate ordibutylsebacate can be used. To this mixture is added talc (50% wt ofdry polymer in the EUDRAGIT™) to form a dispersion. The rapid releasegranules prepared in Example 2 are coated in a fluidized bed with thisdispersion until a 15% wt increase in granule weight is achieved.

[0241] Slow Release.

[0242] A mixture is prepared as follows. EUDRAGIT™ RS 30D (1000 g, 15%wt. aqueous dispersion, AQUA-COAT™ OR SURELEASE) is plasticized withtriethylcitrate (TEC, 20% wt. of dry polymer in the EUDRAGIT™) for 1-24hours. Talc (50% wt. of dry polymer in the EUDRAGIT™ is added withmixing to form the mixture. The rapid release granules prepared inExample 2 are coated with this mixture until a 10-15% wt. increase ingranule weight is achieved. The coated granules are then coated with anEUDRAGIT™ S100 dispersion as done immediately above until a 10-15% wt.increase in granule weight is achieved.

EXAMPLE 9 Preparation of DFMO-Containing Sustained Release Granule

[0243] This procedure employs a double granulation. Thus, DFMO (500 g),MCC (500 g) and EUDRAGIT™ RS 30D (75-100 g) are mixed. To this mixtureis added sufficient water to yield a 30% wt. suspension. To thesuspension is added TEC(10% wt. based on dry polymer weight ofEUDRAGIT™) to yield a dispersion which is wet granulated and dried toremove as much water as possible. The granules are then ground into afine powder. Th the powder is added sufficient water to make a wet masswhich is extruded, spheronized, dried, ground and screened (size 14-20mesh).

EXAMPLE 10 Preparation of DFMO-Containing Capsule Comprising Gastric.Enteric and Colorectal Release Granules

[0244] The following procedure details the preparation of the dosageform described by FIG. 5. Rapid gastric release granules (450 g,prepared according to Example 2), rapid enteric release granules (100 g,prepared according to Example 6) and slow colorectal release granules(450 g, prepared according to Example 8) are mixed thoroughly. Hardgelatin capsules (2000) are then filled with the mixture usingprocedures and equipment well known in the art.

EXAMPLE 11 Preparation of DFMO-Containing Tablets Comprising Rapid andSlow Colorectal Release Granules

[0245] The following procedure details the preparation of the dosageform described by FIG. 12 and comprises a slow release core surroundedby a rapid release layer which is then surrounded by a pH responsivecoating for colorectal release. Thus, DFMO (600 g) is dry blended withAVICEL PH101 (260 g), PVP (30 g), HMPC (100 g, KISM), fumed silicondioxide (50 g, CAB-O-SIL M5P), and magnesium stearate (50 g). Themixture is then compressed into tablets. HPMC (70-80 g, aqueous coat,Opadry®, plasticized with propylene glycol) and DFMO (20-30 g) are mixedand turned into a dispersion by the addition of water. The tablets justprepared are coated with the HPMC/DFMO dispersion until the desiredtablet weight gain has been achieved to form a two layered tablet. Thelayered tablet is then coated in a perforated pan coating unit withEUDRAGIT™ S100 until a 15% wt. increase in tablet weight is achieved.

EXAMPLE 12 Pellet Formulation

[0246] The present example is provided to demonstrate the utility of thepresent invention for the preparation of pellets. TABLE 3 Compound %(w/w) Eflornithine Hydrochloride 50 Lot # A1981-004 Avicel ® PH101 47Kollidon ® K90 3 Total 100

[0247] Procedure:

[0248] a. Kollidon® K90 prepared as a 10% w/w aqueous solution withdistilled water.

[0249] b. Particle size of bulk drug reduced with porcelain mortar andpestle and dry blended for 10 minutes with Avicel® PH 101.

[0250] c. Powder blend transferred to a planetary mixer and theKollidon® K90 solution slowly added.

[0251] d. An additional aliquot of approximately 5% to 13% (based on dryweight of formulation) of distilled water was added to the powder/PVPmass to achieve desirable wet massing.

[0252] e. Final wet mass allowed to mix for an additional 3-5 minutes.

[0253] 1. Extrusion

[0254] Procedure:

[0255] a. Extrusion of the wet mass in Table 3 was accomplished using abench top extruder fitted with a 1.0 mm screen.

[0256] b. The rotation speed was set at maximum (approximately 25 rpm).

[0257] c. The extruded strands were collected periodically and processedfurther.

[0258] 2. Spheronization

[0259] Procedure:

[0260] a. Spheronization was accomplished using a bench top spheronizerfitted with a fine cut plate.

[0261] b. An aliquot of approximately 100-150 gm of extruded materialwas charged into the chamber and spheronized as indicated below.

[0262] c. Periodic dusting of the pellets with Avicel® PH101 was done toabsorb expired moisture produced during the spheronization in order toreduce particle adhesion and limit particle growth.

[0263] d. The wet pellets were collected and gently screened through a14 and 18 mesh screen to remove fines and oversized particles. Theunder/over sized material was recycled into the extrusion mass toincrease yield and decrease waste.

[0264] e. TABLE 4 Spheronization Time/Speed Time (min) Percent ofMaximum 1  50% 1  75% 2-3 100% Total Spheronization Time 4-5 min

[0265] 3. Collection and Sizing

[0266] Procedure:

[0267] a. The wet pellets were collected and evenly spread onto aluminumsheets.

[0268] b. These sheets were then placed in 40° C. ovens, and the pelletswere allowed to dry overnight.

[0269] c. The dried pellets were collected and vacuumed dusted to removeany free particulate matter.

[0270] d. The final bead distribution was assessed using standard metalscreens and is shown below.

[0271] e. The percent of lost material in processing was less than 3%.

[0272] f. Pellets of the 16-18 mesh and 18-20 mesh size were blended andused in subsequent procedures. TABLE 5 Pellet Size Distribution PassThrough (mesh) Retained On (mesh) % of Starting Material 12 16 19.7 1618 54.9 18 20 16.4 20 30  6.2 Total 97.2

[0273] 4. Pellet Coating

[0274] Equipment:

[0275] a. All coatings were applied with using a fluidized bed coaterfitted with a Wurster insert and a bottom spray technique. A filter bagassembly was used, and the insert adjusted from {fraction (3/8)}″ to{fraction (1/2)}″ above the floor of the coating chamber in order toobtain a desirable flow of pellets through the spray path.

[0276] b. Coating material was supplied with a peristalic pump and thespray rate adjusted as indicated.

[0277] c. The atomization pressure was 0.8-1.2 bar, and the filter blowout pressure was set at 1-2 bar.

[0278] d. The coating charge unless otherwise stated was 300 gm of size16-20 mesh pellets as indicated above.

[0279] Enteric Coating TABLE 6 Coating Formulation Material Based OnWeight EUDRAGIT ™  87.00 gm L30D-55 Triethyl Citrate (TEC) 15%, dryweight polymer  3.92 gm Talc 50%, dry weight polymer  13.05 gm WaterTotal solids to be 15% 196.18 gm Total Suspension Weight 300.15 gm

[0280] Calculation:

[0281] For an 8% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.08$

[0282] where x is 26.1 gm and is provided by 87.00 gm of 30% polymersuspension.

[0283] Procedure:

[0284] a. The EUDRAGIT™ L30D-55 and TEC were combined and allowed tostir for 30 minutes.

[0285] b. The talc was separately dispersed in the water for 10 minutesat 4500 rpm.

[0286] c. The talc suspension was added with stirring to theEUDRAGIT™/TEC suspension. TABLE 7 Spray Process Temperature (° C.) AirVolume Time (min) Spray Rate (g/min) In/Out (M³/H) 5 0 45/33 50 15 1.540/32 60 130 2.5 40/30 50-60

[0287] Curing:

[0288] a. Coated beads were collected and spread evenly onto aluminumsheets.

[0289] b. Final product allowed to dry at room temperature overnight.

[0290] Colonic Coating TABLE 8 Coating Formulation Material Based OnWeight EUDRAGIT ™  87.00 gm 4110D Triethyl Citrate (TEC) 5%, dry weightpolymer  1.3 gm  Imwitor ® 900 5%, dry weight polymer  1.3 gm  WaterTotal solids to be 15%  93.1 gm  Total Suspension Weight 182.7 gm 

[0291] Calculation:

[0292] For an 8% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.08$

[0293] where x is 26.1 gm and is provided by 87.00 gm of 30% polymersuspension.

[0294] Procedure:

[0295] a. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rmp for 15 minutes. The resultantdispersion is then allowed to cool to less than 30° C. before adding tothe EUDRAGI™ 4110D.

[0296] b. The final dispersion is allowed to stir for 15 minutes priorto application. TABLE 9 Spray Process Temperature (° C.) Air Volume Time(min) Spray Rate (g/min) In/Out (M³/H) 5 0 30/27 50 15 2.0 33/26 50 652.5 36/26 50

[0297] Curing:

[0298] a. Coated beads were collected and spread evenly onto aluminumsheets.

[0299] b. Final product allowed to dry at room temperature overnight.

[0300] Extended Release Coating TABLE 10 Coating Formulation MaterialBased On Weight EUDRAGIT ™ RS30D 100.00 gm EUDRAGIT ™ RL30D 11.10 gmTriethyl Citrate (TEC) 15%, dry weight polymer 5.00 gm Imwitor ® 900 5%, dry weight polymer 1.67 gm Water Total solids to be 15% 115.40 gmTotal Suspension Weight 233.13 gm

[0301] Calculation:

[0302] For a 10% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.1$

[0303] where x is 33.3 gm and is provided by 111.10 gm of 30% polymersuspension in a ratio of 9:1 RS30D to RL30D.

[0304] Pocedure:

[0305] a. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rpm for 15 minutes. The resultantdispersion is then allowed to cool to less than 30° C. before adding tothe EUDRAGIT™ RS/RL dispersion.

[0306] b. The final dispersion is allowed to stir for 15 minutes priorto application. TABLE 11 Spray Process Time Temperature (° C.) (min)Spray Rate (g/min) In/Out Air Volume (M³/H) 5 0 35/32 25 115 2.1-2.338/29 70 15 0 38/38 70

[0307] Curing:

[0308] 1. Coated beads were dried in situ for 15 minutes with the filterscreen in place in preparation for the next application

[0309] Colonic Coating of Extended Release Pellets TABLE 12 CoatingFormulation Material Based On Weight EUDRAGIT ™ 4110D 95.70 gm TriethylCitrate (TEC) 5%, dry weight polymer 1.40 gm Imwitor ® 900 5%, dryweight polymer 1.40 gm Water Total solids to be 15% 102.20 gm TotalSuspension Weight 200.70 gm

[0310] Calculation:

[0311] For a 8% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.08$

[0312] where x is 28.7 gm and is provided by 95.70 gm of 30% polymersuspension. Here, the coating charge is increased due to the weight gainof solid material from the previous spray application.

[0313] Procedure:

[0314] a. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rmp for 15 minutes. The resultantdispersion is then allowed to cool to less than 30° C. before adding tothe EUDRAGIT™ 4110D dispersion.

[0315] b. The final dispersion is allowed to stir for 15 minutes priorto application. TABLE 13 Spray Process Time Temperature (° C.) (min)Spray Rate (g/min) In/Out Air Volume (M³/H) 5 0 35/30 70 65 2.1-2.538/29 70

[0316] Beads were immediately coated with a 0.2% Imwitor® 900 top coatto prevent sticking during drying and storage.

[0317] 0.2% GMS Top Coat TABLE 14 Coating Formulation Material Based OnWeight Imwitor ® 900 0.70 gm Water Total solids to be 2% 34.30 gm TotalSuspension Weight 35.00 gm

[0318] Calculation:

[0319] For a 0.2% weight gain of polymer (x is dry weight of polymer)$\frac{x}{350 + x} = 0.002$

[0320] where x is 0.7 gm. Here, the coating charge is increased due tothe weight gain of solid material from the previous spray applications.

[0321] Procedure:

[0322] 1. The Imwitor® 900, and water are heated to approximately 65° C.and then homogenized at 4000 rpm for 15 minutes. TABLE 15 Spray ProcessTime Temperature (° C.) (min) Spray Rate (g/min) In/Out Air Volume(M³/H) 3 0 39/33 70 15 2.1-2.5 39/30 70 10 0 40/40 70

[0323] Curing:

[0324] a. Coated beads were collected and spread evenly onto aluminumsheets.

[0325] Final product allowed to dry at room temperature over theweekend.

[0326] Dissolution Testing

[0327] Apparatus

[0328] b. Testing was done using the USP Method II, rotating paddleprocedure.

[0329] c. All testing was done at 37° C. and 100 rpm paddle speed.

[0330] d. Various dissolution media was used to approximate the in vivoconditions that the pellets may experience.

[0331] e. Three milliliter samples were drawn at set time points, andblank dissolution media was added to maintain a constant dissolutionvolume.

[0332] f. Final dissolution volumes were 500 ml except for testing ofthe EUDRAGIT® L30D-55 product where the final volume was 467 ml.

[0333] g. Initial media was 0.1 N HCL and, if needed, aliquots of 0.2 Msodium phosphate tribasic was added to increase the pH. At pH changesafter the addition of the 0.2 M sodium phosphate, the pH of theindividual kettles was adjusted with 2 M HCL or 2 M NaOH prior tosampling.

[0334] h. Dissolution samples were filtered with a 0.45 pm PTFE filterprior to analysis. TABLE 16 Dissolution Time and Media Product TimeMedia pH Uncoated Pellets  0-120 min 0.1 N HCI 1.0 500 ml EUDRAGIT ™ 0-120 min 0.1 N HCI 1.0 L30D055 350 ml 120-165 min +117 ml 6.8 0.2 MNaPO EUDRAGIT ™  0-120 min 0.1 N HCI 1.0 4110D 375 ml 120-240 min +87.5ml 6.0 0.2 M NaPO 240-360 min +37.5 ml 7.4 0.2 M NaPO EUDRAGIT ™  0-120min 0.1 N HCI 1.0 RS/RL w/ 375 ml EUDRAGIT ™ 4110D 120-240 min +87.5 ml6.0 0.2 M NaPO 240-870 min +37.5 ml 7.4 0.2 M NaPO

[0335] 5. Drug Content

[0336] Procedure:

[0337] a. An aliquot of pellets was ground with a porcelain mortar andpestle.

[0338] b. Approximately 500 mg aliquot of the ground material wastransferred to a 250 ml volumetric flask and brought to 3/4 volume withdistilled water.

[0339] c. The flasks were then sonicated for 20 minutes and allowed tocool to room temperature before bringing up to volume with distilledwater.

[0340] d. Sample aliquots were filtered with a 10 pm and then a 0.45 pmfilter prior to analysis.

EXAMPLE 13 Pellet Formulation Rapid Release Core Pellet and EntericPolymer Coated Pellet

[0341] The present example is provided to demonstrate the DFMO releaseprofile of an uncoated rapid release pellet. The dissolution profile ofthis formulation is demonstrated at FIG. 14. TABLE 17 Pellet FormulationCompound % (w/w) Eflornithine Hydrochloride 50 Lot #A1981-004 Avicel ®PH101 47 Kollidon ® K90 3 Total 100

[0342] The particle size distribution of the dried extruded pellets isshown in Table 19. FIG. 14 shows the DFMO is rapidly released, with over90% of the drug being released within 5 minutes when placed in 0.1 NHCl. (uncoated=(

) (see FIG. 14).

[0343] Procedure:

[0344] 1. Kollidon® K90 prepared as a 10% w/w aqueous solution withdistilled water.

[0345] 2. Particle size of bulk drug reduced with porcelain mortar andpestle and dry blended for 10 minutes with Avicel® PH 101.

[0346] 3. Powder blend transferred to a planetary mixer and theKollidon® K90 solution slowly added.

[0347] 4. An additional aliquot of approximately 5% to 13% (based on dryweight of formulation) of distilled water was added to the powder/PVPmass to achieve desirable wet massing.

[0348] 5. Final wet mass allowed to mix for an additional 3-5 minutes.

[0349] Extrusion

[0350] Procedure:

[0351] 1. Extrusion of the wet mass was accomplished using a bench topextruder fitted with a 1.0 mm screen.

[0352] 2. The rotation speed was set at maximum (approximately 25 rpm).

[0353] 3. The extruded strands were collected periodically and processedfurther.

[0354] Spheronization

[0355] 1. Spheronization was accomplished using a bench top spheronizerfitted with a fine cut plate.

[0356] 2. An aliquot of approximately 100-150 gm of extruded materialwas charged into the chamber and spheronized as indicated below.

[0357] 3. Periodic dusting of the pellets with Avicel® PH101 was done toadsorb expired moisture produced during the spheronization in order toreduce particle adhesion and limit particle growth.

[0358] 4. The wet pellets were collected and gently screened through a14 and 18 mesh screen to remove fines and oversized particles. Theunder/over sized material was recycled into the extrusion mass toincrease yield and decrease waste. TABLE 18 Spheronization Time/SpeedTime (min) Percent of Maximum 1 50% 1 75% 2-3 100% Total SpheronizationTime 4-5 min

[0359] Collection and Sizing

[0360] Procedure:

[0361] 1. The wet pellets were collected and evenly spread onto aluminumsheets.

[0362] 2. These sheets were then placed in 40° C. ovens, and the pelletswere allowed to dry overnight.

[0363] 3. The dried pellets were collected and vacuumed dusted to removeany free particulate matter.

[0364] 4. The final bead distribution was assessed using standard metalscreens and is shown below.

[0365] 5. The percent of lost material in processing was less than 3%.

[0366] 6. Pellets of the 16-18 mesh and 18-20 mesh size were blended andused in subsequent procedures. TABLE 19 Pellet Size Distribution PassThrough (mesh) Retained On (Mesh) % of Starting Material 12 16 19.7 1618 54.9 18 20 16.4 20 30 6.2 Total 97.2

[0367] The following formula was employed to prepare the pellets coatedwith an enteric polymer of the present invention.

[0368] As demonstrated in FIG. 15, release of the active ingredient DFMOwas less than 10% after 1 hour and less than 20% after 2 hours. Theserelease rates could be decreased with higher levels of polymer beingapplied to the pellets.

[0369] The profile mirrors the delayed plasma level drug (DFMO)illustrated in FIG. 1, middle profile (Rapid enteric release).

[0370] Pellet Coating

[0371] Equipment:

[0372] 1. All coatings were applied with using a fluidized bed coaterfitted with a Wurster insert and a bottom spray technique.

[0373] 2. Coating material was supplied with a peristaltic pump and thespray rate adjusted as indicated.

[0374] 3. The atomization pressure was 0.8-1.2 bar, and the filter blowout pressure was set at 1-2 bar.

[0375] 4. The coating charge unless otherwise stated was 300 gm of size16-20 mesh pellets as indicated above.

[0376] Enteric Coating TABLE 20 Coating Formulation Material Based OnWeight EUDRAGIT ™ L30D-55  87.00 gm Triethyl Citrate (TEC) 15%, dryweight polymer  3.92 gm Talc 50%, dry weight polymer  13.05 gm WaterTotal solids to be 15% 196.18 gm Total Suspension Weight 300.15 gm

[0377] Calculation:

[0378] For an 8% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.08$

[0379] where x is 26.1 gm and is provided by 87.00 gm of 30% polymersuspension.

[0380] Procedure:

[0381] 1. The EUDRAGIT™ L30D-55 and TEC were combined and allowed tostir for 30 minutes.

[0382] 2. The talc was separately dispersed in the water for 10 minutesat 4500 rpm.

[0383] 3. The talc suspension was added with stirring to theEUDRAGIT™/TEC suspension. TABLE 21 Spray Process Time Temperature (° C.)(min) Spray Rate (g/min) In/Out Air Volume (M³/H) 5 0 45/33 50 15 1.540/32 60 130 2.5 40/30 50-60

[0384] Curing:

[0385] 1. Coated beads were collected and spread evenly onto aluminumsheets.

[0386] 2. Final product allowed to dry at room temperature overnight.

[0387]FIG. 15 illustrates that with 8% weight gain of EUDRAGIT™ L30D-55,less than 20% of the drug was released in acidic medium. Rapid drugrelease was evident when the medium was changed to pH 6.8.

[0388] Colonic Coating TABLE 22 Coating Formulation Material Based OnWeight EUDRAGIT ™ 4110D 87.00 gm Triethyl Citrate (TEC) 5%, dry weightpolymer 1.3 gm Imwitor ® 900 5%, dry weight polymer 1.3 gm Water Totalsolids to be 15% 93.1 gm Total Suspension Weight 182.7 gm

[0389] Calculation:

[0390] For an 8% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.08$

[0391] where x is 26.1 gm and is provided by 87.00 gm of 30% polymersuspension.

[0392] Procedure:

[0393] 1. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rpm for 15 minutes. The resultantdispersion is then allowed to cool to less than 30° C. before adding tothe EUDRAGIT™ 4110D.

[0394] 2. The final dispersion is allowed to stir for 15 minutes priorto application. TABLE 23 Spray Process Time Temperature (° C.) (min)Spray Rate (g/min) In/Out Air Volume (M³/H) 5 0 30/27 50 15 2.0 33/26 5065 2.5 36/26 50

[0395] Curing:

[0396] 1. Coated beads were collected and spread evenly onto aluminumsheets.

[0397] 2. Final product allowed to dry at room temperature overnight.

[0398] The DFMO release profile of the colonic coating preparation(third peak) (-∘-∘-∘-) is further illustrated in FIG. 16. In FIG. 16 itis demonstrated that less than 20% of the drug was released after 4hours when the coated pellets were first subjected to pH media 1.0 for 2hours, and then followed by pH media 6.0 for 2 hours. Since EUDRAGIT™4110D starts to dissolve at pH 6.8, rapid release of the drug was seenwhen the pH of the media was adjusted to pH 7.4. The anticipated plasmablood level profile is represented in the far right profile of FIG. 1(open circles, -∘-∘-∘-).

EXAMPLE 14 Extended Release Coating

[0399] To extend the release profile of this very water-soluble drug(DFMO) in the colon, an extended release coating was applied to thepellets. TABLE 24 Extended Release Coating Formulation Material Based OnWeight EUDRAGIT ™ RS30 100.00 gm EUDRAGIT ™ RL30D  11.10 gm TriethylCitrate (TEC) 15%, dry weight polymer  5.00 gm Imwitor ® 900  5%, dryweight polymer  1.67 gm Water Total solids to be 15% 115.40 gm TotalSuspension Weight 233.13 gm

[0400] Calculation:

[0401] For a 10% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.1$

[0402] where x is 33.3 gm and is provided by 111.10 gm of 30% polymersuspension in a ratio of 9:1 RS30D to RL30D.

[0403] Procedure:

[0404] 1. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rpm for 15 minutes. The resultantdispersion is then allowed to cool to less than 30° C. before adding tothe EUDRAGIT™ RS/RL dispersion. TABLE 25 Spray Process Time Temperature(° C.) (min) Spray Rate (g/min) In/Out Air Volume (M³/H) 5 0 35/32 25115 2.1-2.3 38/29 70 15 0 38/38 70

[0405] Curing:

[0406] 1. Coated beads were dried in situ for 15 minutes with the filterscreen in place in preparation for the next application.

[0407] Colonic Coating of Extended Release Pellets:

[0408] Coat #2 (outside coat) TABLE 26 Coating Formulation MaterialBased On Weight EUDRAGIT ™ 4110D  95.70 gm Triethyl Citrate (TEC) 5%,dry weight polymer  1.40 gm Imwitor ® 900 5%, dry weight polymer  1.40gm Water Total solids to be 15% 102.20 gm Total Suspension Weight 200.70gm

[0409] This coat protects the pellet in the GI tract until a pH ofgreater than 6.8 is achieved. At that pH and above, the 4110D begins todissolve.

[0410] Calculation:

[0411] For an 8% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.08$

[0412] where x is 28.7 gm and is provided by 95.70 gm of 30% polymersuspension. Here, the coating charge is increased due to the weight gainof solid material from the previous spray application.

[0413] Procedure:

[0414] 1. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rpm for 15 minutes. The resultantdispersion is then allowed to cool to less than 30C before adding to theEUDRAGIT™ 4110D dispersion.

[0415] 2. The final dispersion is allowed to stir for 15 minutes priorto application. TABLE 27 Spray Process Time Temperature (° C.) (min)Spray Rate (g/min) In/Out Air Volume (M³/H) 5 0 35/30 70 65 2.1-2.538/29 70

[0416] Beads were immediately coated with a 0.2% Imwitor® 900 top coatto prevent sticking during drying and storage.

[0417] 0.2% GMS Top Coat TABLE 28 Coating Formulation Material Based OnWeight Imwitor ® 900  0.70 gm Water Total solids to be 2% 34.30 gm TotalSuspension Weight 35.00 gm

[0418] Calculation:

[0419] For a 0.2% weight gain of polymer (x is dry weight of polymer)$\frac{x}{300 + x} = 0.002$

[0420] where x is 0.7 gm. Here, the coating charge is increased due tothe weight gain of solid material from the previous spray applications.

[0421] Procedure:

[0422] 1. The Imwitor® 900, and water are heated to approximately 65° C.and then homogenized at 4000 rpm for 15 minutes. TABLE 29 Spray ProcessTime Temperature (° C.) (min) Spray Rate (g/min) In/Out Air Volume(M³/H) 3 0 39/33 70 15 2.1-2.5 39/30 70 10 0 40/40 70

[0423] Curing:

[0424] 1. Coated beads were collected and spread evenly onto aluminumsheets.

[0425] 2. Final product allowed to dry at room temperature over theweekend.

[0426] As shown in FIG. 17, drug release from the extended releasepellet formulations coated with EUDRAGIT™ 4110D did not release drugduring the first 5 hours that the pellets were exposed to pH 1.0, 6.0,and then 7.0 after 7 hours, approximately 17% of the drug was releasedand after about 14½ hours, approximately 55% of the drug was releasedfrom the pellets. In FIG. 2, a pellet coated with a combination of RS30and RL30D and a top coat of 4110D (-⋄-⋄-⋄-) is represented in anexpected in vivo blood release profile of DFMO. A rapid release pelletcoated with 4110D is illustrated in FIG. 2, open circles. The subcoat ofthe various pharmaceutical preparations described herein may includeother materials with properties of EUDRAGIT™ 4110D, that would producesimiliar results. Such would include materials that dissolve ordisintegrate above pH 6.8. By example, these are EUDRAGIT™ S100 or HPMCacetate succinate derivative, and polyanhydrides and other retardantmaterials that are broken down or degraded by enzymes and bacteria inthe colon.

[0427] Dissolution Testing

[0428] Apparatus

[0429] 1. Testing was done using the USP Method II, rotating paddleprocedure.

[0430] 2. All testing was done at 37° C. and 100 rpm paddle speed.

[0431] 3. Various dissolution media was used to approximate the in vivoconditions that the pellets may experience.

[0432] 4. Three milliliter samples were drawn at set time points, andblank dissolution media was added to maintain a constant dissolutionvolume

[0433] 5. Final dissolution volumes were 500 ml except for testing ofthe EUDRAGIT™ L30D-55 product where the final volume was 467 ml.

[0434] 6. Initial media was 0.1 N HCL and, if needed, aliquots of 0.2 Msodium phosphate tribasic was added to increase the pH. At pH changesafter the addition of the 0.2 M sodium phosphate, the pH of theindividual kettles was adjusted with 2 Mm HCL or 2 M NaOH prior tosampling. TABLE 30 Dissolution Time and Media Product Time Media pHUncoated Pellets  0-120 min 0.1 N HCl 1.0 500 ml EUDRAGIT ™  0-120 min0.1 N HCl 1.0 L30D-55 350 ml 120-165 min +117 ml 6.8 0.2 M NaPO₄EUDRAGIT ™  0-120 min 0.1 N HCl 1.0 4110D 375 ml 120-240 min +87.5 ml6.0 0.2 M NaPO₄ 240-360 min +37.5 ml 7.4 0.2 M NaPO₄ EUDRAGIT ™  0-120min 0.1 N HCl 1.0 RS/RL w/ 375 ml EUDRAGIT ™ 4110D 120-240 min +87.5 ml6.0 0.2 M NaPO₄ 240-870 min +37.5 ml 7.4 0.2 M NaPO₄

[0435] Drug Content

[0436] Procedure:

[0437] 1. An aliquot of pellets was ground with a porcelain mortar andpestle.

[0438] 2. Approximately 500 mg aliquot of the ground material wastransferred to a 250 ml volumetric flask and brought to {fraction (3/4)}volume with distilled water.

[0439] 3. The flasks were then sonicated for 20 minutes and allowed tocool to room temperature before bringing up to volume with distilledwater.

[0440] 4. Sample aliquots were filtered with a 10 μm and then a 0.45 μmfilter prior to analysis.

[0441] Drug Analysis HPLC Conditions Column: Rainin Microsorb ™Short-One ® C18, ODS-1, 3 μm Mobile Phase: 1.1 mM Sodium Dodecyl Sulfatein 23/77 Acetonitrile/ 0.039 M NaPO₄ (pH = 2.3) Flow Rate: 1.0 ml/minDetection: 210 nm Injection Volume: 50 μl Retention Time: 4.5 minutes

EXAMPLE 15 DFMO-Containing Slow Release Matrix Pellets—No Coating

[0442] The present example is provided to demonstrate the utility of thepresent formulation in providing a slow release matrix pelletformulation of DFMO.

[0443] I. Pellet Formulation TABLE 31 Compound % (w/w) EflornithineHydrochloride 46.9 Lot #R41063 Avicel ® RC591 10.4 Sterotex K 26.1EUDRAGIT ™ NE40D (dry polymer) 16.6 Total 100

[0444] Procedure:

[0445] 1. Aggregates of bulk drug were reduced with a porcelain mortarand pestle and dry blended for 5 minutes with Avicel® RC591 and SterotexK.

[0446] 2. Powder blend transferred to a planetary mixer, and theEUDRAGIT™ NE40D dispersion was slowly added.

[0447] 3. An additional aliquot of approximately 10% to 15% (based ondry weight of formulation) of distilled water was added to thepowder/EUDRAGIT™ NE40D mass to achieve desirable wet massing.

[0448] 4. Final wet mass allowed to mix for an additional 3-5 minutes.

[0449] II. Extrusion

[0450] Procedure:

[0451] 1. Extrusion of the wet mass in (I) was accomplished using abench top extruder fitted with a 1.0 mm screen.

[0452] 2. The rotation speed was set at maximum (approximately 25 rpm).

[0453] 3. The extruded strands were collected periodically and processedfurther.

[0454] III. Spheronization

[0455] 1. Spheronization was accomplished using a bench top spheronizerfitted with a fine cut plate.

[0456] 2. An aliquot of approximately 100-150 gm of extruded materialwas charged into the chamber and spheronized as indicated below.

[0457] 3. An air assist pressure of 0.2 Mpa was used to decrease thetackiness of the beads during spheronization.

[0458] 4. The wet pellets were collected and gently screened through a14 mesh screen to remove oversized particles. The oversized material wasrecycled into the extrusion mass to increase yield and decrease waste.TABLE 32 Spheronization Time/Speed Time (min) Percent of Maximum 3-3.580% Total Spheronization Time 3-4 min

[0459] IV. Collection and Sizing

[0460] Procedure:

[0461] 1. The wet pellets were collected and evenly spread onto aluminumsheets.

[0462] 2. These sheets were then left at room temperature to dryovernight.

[0463] 3. The dried pellets were collected, and the final beaddistribution was assessed using standard metal screens and is shownbelow.

[0464] 5. The percent of lost material in processing was less than 8%.

[0465] 6. Pellets of the 16-20 mesh size were used in subsequentprocedures. TABLE 33 Pellet Size Distribution Pass Through (mesh)Retained On (Mesh) % of Starting Material 12 16 15.8 16 20 60.7 20Collection Plate 15.6 Total 92.1

[0466] V. Matrix Melting

[0467] Equipment:

[0468] 1. Melting of the wax matrix was accomplished with a fluidizedbed coater fitted with an acrylic coating chamber and a filter screen.

[0469] 2. The pellet cores were heated at 65° C. for 15 minutes and thenallowed to cool at room temperature on aluminum sheets.

[0470] The dissolution release profile of the uncoated slow-releaseDFMO-containing matrix pellets are illustrated at FIG. 18. In FIG. 18(----) the dissoltion curve of DFMO from the matrix pellet is 0.1 NHcl. The slight decrease in DFMO release in pH 7.4 is due to therelatively lower solubility of the drug DFMO at a neutral pH.

EXAMPLE 16 Release of DFMO from Coated Matrix Beads

[0471] The present example illustrates the use of the present inventionwith coated matrix beads containing DFMO. The release profiles here inan acidic media reflect poor adhesion of the EUDRAGIT™ L30D-55 andEUDRAGIT™ 4110D to the wax containing pellets containing DFMO. The rapidrelease of the drug was evident from both pellet formulations in theacidic medium as demonstrated in FIG. 19.

[0472] VI. Pellet Coating

[0473] Equipment.

[0474] 1. All coatings were applied with using a fluidized bed coaterfitted with a Wurster insert and a bottom spray technique. A filter bagassembly was used, and the insert adjusted to {fraction (1/2)}″ abovethe floor of the coating chamber in order to obtain a desirable flow ofpellets through the spray path.

[0475] 2. Coating material was supplied with a peristaltic pump, and thespray rate adjusted as indicated.

[0476] 3. The atomization pressure was 1.0-1.2 bar, and the filter blowout pressure was set at 2 bar.

[0477] 4. The coating charge was 300 gm of size 16-20 mesh pellets asdescribed above except in the ethylcellulose/4110D coated product. Theethylcellulose/4110D coated product utilizes 300 gm of the rapid releasebeads detailed in the August 1997 report (lot #07219701).

[0478] Enteric Coating TABLE 34 Coating Formulation Material Based OnWeight EUDRAGIT ™ L30D-55 88.9 gm Triethyl Citrate (TEC) 15%, dry weightpolymer 4.00 gm Imwitor ® 900  5%, dry weight polymer 1.3 gm Water Totalsolids to be 15% 92.5 gm Total Suspension Weight 186.7 gm

[0479] Calculation:

[0480] For an 8.2% weight gain of polymer (x is dry weight of polymer)x/(300+x)=0.08 where x is 26.7 gm and is provided by 88.9 gm of 30%polymer suspension.

[0481] Procedure:

[0482] 1. The TEC, Imwitor® 90, and water are heated to approximately65° C. and then homogenized at 4000 rpm for 15 minutes. The resultantdispersion is then allowed to cool to less than 30° C. before adding tothe EUDRAGIT™ L30D-55.

[0483] 2. The final coating dispersion is allowed to mix for at least 30minutes prior to application. TABLE 35 Spray Process Time Temperature (°C.) (min) Spray Rate (g/min) In/Out Air Volume (M³/H) 5 0 40/31 65 202.4 40/32 66 80 2.5 41/30 70

[0484] Beads were immediately coated with a 0.2% Imwitor® 900 top coatto prevent sticking during drying and storage.

[0485] 0.2% GMS Top Coat TABLE 36 Coating Formulation Material Based OnWeight Imwitor ® 900  0.66 gm Water Total solids to be 2% 32.54 gm TotalSuspension Weight 33.20 gm

[0486] Calculation:

[0487] For an 0.2% weight gain of polymer (x is dry weight of polymer)x/(332+x)=0.002 where x is 0.66 gm. Here, the coating charge isincreased due to the weight gain of solid material from the previousspray applications.

[0488] Procedure:

[0489] 1. The Imwitor® 900 and water are heated to approximately 65° C.and then homogenized at 4000 rpm for 15 minutes. TABLE 37 Spray ProcessTime Temperature (° C.) (min) Spray Rate (g/min) In/Out Air Volume(M³/H) 5 0 43/33 70 15 2.0 40/32 70 25 0 40/40 70

[0490] Curing:

[0491] 1. Coated beads were collected and spread evenly onto aluminumsheets.

[0492] 2. Final product was cured at 40° C. for 2 hours and then at roomtemperature overnight.

[0493] Colonic Coating TABLE 38 Coating Formulation Material Based OnWeight EUDRAGIT ™ 4110D  88.9 gm Triethyl Citrate (TEC) 5%, dry weightpolymer  1.3 gm Imwitor ® 900 5%, dry weight polymer  1.3 gm Water Totalsolids to be 15%  95.2 gm Total Suspension Weight 186.7 gm

[0494] Calculation:

[0495] For an 8.2% weight gain of polymer (x is dry weight of polymer)x/(300+x)=0.08 where x is 26.7 gm and is provided by 88.9 gm of 30%polymer suspension.

[0496] Procedure:

[0497] 1. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rpm for 15 minutes. The resultantdispersion is then allowed to cool to less than 30° C. before adding tothe EUDRAGIT™ 4110D.

[0498] 2. The final dispersion is allowed to mix for at least 30 minutesprior to application. TABLE 39 Spray Process Temperature (° C.) AirVolume Time (min) Spray Rate (g/min) In/Out (M³/H) 5 0 35/33 70 15 2.036/29 70 65 2.5 37/26 70

[0499] Beads were immediately coated with a 0.2% Imwitor® 900 top coatto prevent sticking during drying and storage.

[0500] 0.2% GMS Top Coat TABLE 40 Coating Formulation Material Based OnWeight Imwitor ® 900  0.66 gm Water Total solids to be 2% 32.34 gm TotalSuspension Weight 33.00 gm

[0501] Calculation:

[0502] For an 0.2% weight gain of polymer (x is dry weight of polymer)x/(329+x)=0.02 where x is 0.66 gm. Here, the coating charge is increaseddue to the weight gain of solid material from the previous sprayapplications.

[0503] Procedure:

[0504] 1. The Imwitor® 900 and water are heated to approximately 65° C.and then homogenized at 4000 rpm for 15 minutes. TABLE 41 Spray ProcessTemperature (° C.) Air Volume Time (min) Spray Rate (g/min) In/Out(M³/H) 5 0 43/33 70 15 2.0 40/32 70 25 0 40/40 70

[0505] Curing:

[0506] 1. Coated beads were collected and spread evenly onto aluminumsheets.

[0507] 2. Final product was allowed to dry at room temperatureovernight.

[0508] The poor adhesion of the EUDRAGIT™ used (L30D-55 and 4110D) tothe wax containing pellet resulted in poor protection of the core whenexposed to an acidic medium. When the pellets are coated with a 2% to 3%weight gain of Opadry® II prior to coating with the EUDRAGIT, protectionof the pellets to the acidic medium will be provided. The EUDRAGIT™polymers used here demonstrated enhanced adhesion to the Opadry®subcoat. The dissolution profiles of these pellet formulationsrepresented by the two lower profiles in FIG. 19 provide the expectedprofile where a subcoat of Opadry® II is included.

EXAMPLE 17 Extended Release Coating

[0509] These pellets did not include a sub-coating. They were coatedwith AQUA COAT® containing the Methacel, a coating of EUDRAGIT™ 4110Dwas then applied. TABLE 42 Coating Formulation Material Based On WeightAQUA-COAT ™ ® 111.11 gm ECD Methocel ® K4M  3.33 gm Triethyl Citrate(TEC) 20%, dry weight polymer  6.00 gm AQUA-COAT ™ ® ECD Water Totalsolids to be 10% w/w 212.86 gm Total Suspension Weight 333.30 gm

[0510] (See FIG. 20 for data)

[0511] Calculation:

[0512] For a 10% weight gain of polymer (x is dry weight of polymer)x/(300+x)=0.10 where x is 33.3 gm and is provided by 111.10 gm of 27%aqueous ethylcellulose polymer suspension and hydroxypropylmethylcellulose in a ratio of 9:1.

[0513] Procedure:

[0514] 1. The TEC and AQUA-COAT™® -ECD were allowed to mix for at leastthirty minutes before combining with the HPMC solution.

[0515] 2. The HPMC was dispersed in hot water ({fraction (1/3)} of waterfor addition) and then diluted ({fraction (2/3)} of water for addition).The HPMC was allowed to hydrate and form a cool solution prior toaddition to the AQUA-COAT™® ECD dispersion.

[0516] 3. The final dispersion was stirred for at least 30 minutes priorto application. TABLE 43 Spray Process Temperature (° C.) Air VolumeTime (min) Spray Rate (g/min) In/Out (M³/H) 5 0 60/35 70 140 2.1-2.460/44 70 15 0 60/44 70

[0517] Curing:

[0518] 1. Coated beads were dried in situ for 15 minutes with the filterscreen in place in preparation for the next application.

[0519] Colonic Coating of Extended Release Pellets TABLE 44 CoatingFormulation Material Based On Weight EUDRAGIT ™ 4110D  90.49 gm TriethylCitrate (TEC) 5%, dry weight polymer  1.36 gm Imwitor ® 900 5%, dryweight polymer  1.36 gm Water Total solids to be 15%  98.80 gm TotalSuspension Weight 191.01 gm

[0520] Calculation:

[0521] For an 7.4% weight gain of polymer (x is dry weight of polymer)x/(339+x)=0.074 where x is 27.15 gm and is provided by 90.49 gm of 30%polymer suspension. Here, the coating charge is increased due to theweight gain of solid material from the previous spray application.

[0522] Procedure:

[0523] 1. The TEC, Imwitor® 900, and water are heated to approximately65° C. and then homogenized at 4000 rpm for 15 minutes. The resultantdispersion is then allowed to cool to less than 30C before adding to theEUDRAGIT™ 4110D dispersion.

[0524] 2. The final dispersion is allowed to stir for at least 30minutes prior to application. TABLE 45 Spray Process Temperature (° C.)Air Volume Time (min) Spray Rate (g/min) In/Out (M³/H) 5 0 50/41 70 632.1-2.5 39/29 70

[0525] Beads were immediately coated with a 0.2% Imwitor® 900 top coatto prevent sticking during drying and storage.

[0526] 0.2% GMS Top Coat TABLE 46 Coating Formulation Material Based OnWeight Imwitor ® 900  0.74 gm Water Total solids to be 2% 36.16 gm TotalSuspension Weight 36.84 gm

[0527] Calculation:

[0528] For an 0.2% weight gain of polymer (x is dry weight of polymer)x/(369+x)=0.002 where x is 0.74 gm. Here, the coating charge isincreased due to the weight gain of solid material from the previousspray applications.

[0529] Procedure:

[0530] 1. The Imwitor® 900 and water are heated to approximately 65° C.and then homogenized at 4000 rpm for 15 minutes. TABLE 47 Spray ProcessTemperature (° C.) Air Volume Time (min) Spray Rate (g/min) In/Out(M³/H) 3 0 39/33 70 15 2.1-2.5 39/30 70 10 0 40/40 70

[0531] Curing:

[0532] 1. Coated beads were collected and spread evenly onto aluminumsheets.

[0533] 2. Final product dried at 40° C. overnight.

[0534] VI. Dissolution Testing

[0535] Apparatus

[0536] 1. Testing was done using the USP Method I, rotating basketprocedure.

[0537] 2. All testing was done at 37° C. and 100 rpm shaft speed.

[0538] 3. Various dissolution media was used to approximate the in vivoconditions that the pellets may experience.

[0539] 4. Three milliliter samples were drawn at set time points, andblank dissolution media was added to maintain a constant dissolutionvolume

[0540] 5. Final dissolution volumes were 500 ml.

[0541] 6. Initial media was 0.1 N HCL and, if needed, aliquots of 0.2 Msodium phosphate tribasic was added to increase the pH. At pH changesafter the addition of the 0.2 M sodium phosphate, the pH of theindividual kettles was adjusted with 2 N NaOH prior to sampling.

[0542] 7. Dissolution samples were filtered with a 0.45 μm PTFE filterprior to analysis. TABLE 48 Dissolution Time and Media Product TimeMedia pH Uncoated Pellets  0-180 min 0.1 N HCl 1.0 0.05 M Phosphate 7.4500 ml EUDRAGIT ™  0-120 min 0.1 N HCl 1.0 L30D-55 375 ml 120-240 min+125 ml 6.8 0.2 M NaPO₄ EUDRAGIT ™  0-120 min 0.1 N HCl 1.0 4110D 375 ml120-240 min +125 ml 6.8 0.2 M NaPO₄ AQUA-COAT ™ ®  0-120 min 0.1 N HCl1.0 ECD + 375 ml EUDRAGIT ™ 4110D 120-240 min +125 ml 6.8 0.2 M NaPO₄240-480 min Adjust to pH 7.4

[0543] As demonstrated in FIG. 20, retardation of drug release in anacidic (0.1 N HCl) was attained at pH 6.8, the EURDRAGIT™ 4110Ddissolved and during the next 2 hours, the drug diffused through thepores in the AQUA-COAT™ film.

[0544] VII. Drug Content

[0545] Procedure:

[0546] 1. An aliquot of pellets was ground with a porcelain mortar andpestle.

[0547] 2. Approximately 500 mg aliquot of the ground material wastransferred to a 250 ml volumetric flask and brought to {fraction (3/4)}volume with 0.1 N HCl.

[0548] 3. The flasks were then sonicated for 20 minutes and allowed tocool to room temperature before bringing up to volume with distilledwater.

[0549] 4. Sample aliquots were filtered with a 0.45 μm filter prior toanalysis.

[0550] VIII. Drug Analysis HPLC Conditions Column: Alltech PlatinumEPS ™ C18, ODS-1, 5 μm Mobile Phase: 1.1 mM Sodium Dodecyl Sulfate in20/80 v/v Acetonitrile/0.05 M NaPO₄ (pH = 2.3) Flow Rate: 1.0 ml/minDetection: 210 nm Injection Volume: 50 μl Retention Time: 4.2 minutes

EXAMPLE 18

[0551] Multi-layered Matrix Tablet

[0552] The present example is provided to demonstrate the utility of thepresent invention as a multi-layered tablet that includes a core of amatrix containing DFMO, and a slow release material. Such slow releasematerials include by way of example, Klucel™, HPMC, carbomer, Polyox,and other cellulosic and hydrophilic polymers, proteins andpolysaccharides.

[0553] In some embodiments, the tablet will include a core pellet ofDFMO in a matrix. An exemplary matrix retardant used in the presentexample is Klucel. This core tablet will then have a first coat of acolonic protective material, such as EUDRAGIT™ S100. This first coat isthen covered with an Opadry® solution of HPMC or similar rapidlydissolving or freely permeable material, also containing DFMO (2ndcoat). Over this, a third coat of an acid resistant polymer (e.g.EUDRAGIT™ L, CAP, CAT, PVAP, HPMCP, HPMCAS, or similar enteric polymeris applied over the second coat. Then, a fourth coat of an immediaterelease material (e.g. Opadry®) containing DFMO is applied.

[0554] The matrix tablet may be prepared by blending the DFMO (45%) withKlucel HF (25%), spray dried lactose (24%) and Starch 1500 (5%) in asuitable blender for 10 minutes. The magnesium stearate (0.5%) andCabosil M5P (0.5%) are added to the powders. This is blended for anadditional 5 minutes. The powder blend is then compressed into tabletsin the 8-10 kg range in the 200-400 mg range. The tablets are thencoated as described above. Exemplary Weight Gain for Pellet CoatingsCoat 1 EUDRAGIT ™ S100 8% weight gain Coat 3 L30 D-55 8% weight gainCoat 2, Coat 4 HPMC E5:DFMO 10% weight gain  (ratio 5:1)

[0555] Other chemical synthetic techniques well known to those of skillin the art may be used such as that described in the attached list ofreferences which are hereby incorporated in their entirety.

[0556] The above is a detailed description of a particular embodiment ofthe invention. It is recognized that departures from the disclosedembodiment may be made within the scope of the invention and thatobvious modifications will occur to a person skilled in the art. Thefull scope of the invention is set out in the claims that follow andtheir equivalents. Accordingly, the claims and specification should notbe construed to unduly narrow the full scope of protection to which theinvention is entitled.

[0557] Those of skill in the art should, in light of the presentdisclosure, appreciate that many changes can be made in the specificembodiments where are disclosed herein and still obtain a like orsimilar result without departing from the spirit and scope of theinvention. All of the compositions and methods disclosed and claimedherein can be made and executed without undue experimentation in lightof the present disclosure. It will be apparent that certain compoundswhich are both physiologically and chemically related may be substitutedfor the therapeutic compound described herein while the same or similarresults are achieved.

REFERENCES

[0558] The following references, to the extent that they provideexemplary procedural or other details supplementary to those set forthherein, are specifically incorporated herein by reference.

[0559] S. G. Eckhardt, D. Dai, K. K. Davidson, B. J. Forseth, G. M.Wahl, D. D. Von Hoff, Proc. Nat'l. Acad. Sci., USA, 1994, 91, 6674-6679.

[0560] Controlled Drug Delivery: Fundamentals and Applications, 2nd ed.(Joseph R. Robinson and Vincent H. L. Lee, eds., Marcel Dekker, Inc.,NY; 1987) ISBN 0-8247-7588-0

[0561] Aqueous Polymeric Coatings for Pharmaceutical Dosage Forms (JamesW. McGinity, ed.; Marcel Dekker, Inc., NY; 1989) ISBN 0-8247-7907-X

[0562] Pharmaceutical Dosage Forms: Tablets, Vol. 3 (Herbert A.Lieberman, Leon Lachman and Joseph B. Schwartz, eds.; Marcel Dekker,Inc., NY; 1990) ISBN 0-8247-8300-X

[0563] Pharmaceutical Dosage Forms and Drug Delivery Systems, 6th Ed.(Howard C. Ansel, Nicholas G. Popovich and Lloyd V. Allen, Jr., eds.;Williams & Wilkins, Baltimore; 1995) ISBN 0-683-00193-0

[0564] Development of Difluoromethylornithine as a Chemoprevention Agentfor the Management of Colon Cancer, Meyskens, Frank L., Jr.; Gemer,Eugene W. J., J. Cell. Biochem. (1995) (Suppl. 22), 126-31

[0565] Dose De-Escalation Chemoprevention Trial ofAlpha-Difluoromethylomithine in Patients with Colon Polyps. (Meyakens,F. L., Jr; Emerson, S. S; Pelot, D.; Meshkinpour, H.; Shassetz, L. R.;Einspahr J; Alberts, D. S,; Gerner, E. W. 1994 Aug 3) Journal of theNational Cancer Institute, (1994 Aug 3) 86 (15) 1122-30.

[0566] Cancer Chemoprevention. (Lippman S. M.; Benner S. E.; Waun KiHong J. Clin. Oncol., (1994) 12/4 (851-873).

[0567] Alpha-Difluoromethylornithine (DFMO) as a PotentialChemopreventive Agent: Toxicology, Pharmacokinetics and Pharmacodynamicsof Chronic Oral Administration in Humans (Meeting abstract); Creaven, P.J.; Pendyala, L.; Porter, C. W.; Murphy, M. J. Non-serial, (1993).CCPC-93: Second International Cancer Chemo Prevention Conference. April28-30, 1993, Berlin, Germany, p. 53.

[0568] Randomized Phase I Chemoprevention Dose-Seeking Study ofAlpha-Difluoromethylornithine; Love, R. R.; Carbone, P. P.; Verma, A.K.; Gilmore, D.; Carey, P.; Tutsch, K. D.; Pomplun, M.; Wilding, G.,Journal of the National Cancer Institute, (1993 May 5), 85 (9) 732-7.

[0569] Urinary and Erythrocyte Polyamines During the Evaluation of OralAlpha-Difluoromethylornithine in a Phase I Chemoprevention ClinicalTrial; Pendyala, L.; Creaven, P. J.; Porter, C. W., CANCER EPIDEMIOLOGY,BIOMARKERS AND PREVENTION, (1993 May-June) pg. 235-41.

[0570] Polyamine Depletion as a Strategy for Cancer Chemoprevention:Rationale, Problems and Solutions (Meeting Abstract). (Gemer, E. W.Non-serial, Jun. 3-6, 1992, Tucson, Ariz.). Fourth InternationalConference on Prevention of Human Cancer: Nutrition and ChemopreventionControversies.

[0571] PHASE I STUDY OF DIFLUOROMETHYLORNITHINE DFMO AS ACHEMOPREVENTIVE AGENT (CPA) (MEETING ABSTRACT); Creaven, P. J.;Pendyala, L.; Petrelli, N.; Douglass, H.; Herrera, L.; Porter, C.;Solomon, J.; Proc. Annul Meet. Am. Soc. Clin. Oncol. (1992) Vol. 11, pp.A395.

[0572] Polyamine Contents in Rectal and Buccal Mucosae in Humans TreatedWith Oral Difluoromethylornithine; Boyle, J. O.; Meyakens, .F L., Jr.;Garewal, H. S.; Gemer, E. W.; Cancer Epidemiology Biomarkers &Prevention (1992) 1 (2). pg. 131-135.

[0573] Chemoprevention Trials (Meeting Abstract); Kelloff, G. J.;Malone, W. F.; Steele, V.; Boone, C. W.; Third International Conferenceon Mechanisms of Antimutagenesis and Anticarcinogenesis. (May 5-10,1991) Lucca, Italy, p. 42.

[0574] DIFLUOROMETHYLORNITHINE (DFMO), A POTENTIAL CHEMOPREVENTIVE(MEETING ABSTRACT); Carbone, P. P.; Love, R. R.; Carey, P.; Tutsch, K.;Verma, A. K.; Wilding, G.; Gilmore-Cunningham, D.; Proc. Annul Meet. Am.Assoc. Cancer Res., (1991) Vol. 32, pp. A1209.

[0575] Phase I Trial and Pharmacokinetic Study of Intravenous and OralAlpha-Difluoromethylornithine; Griffin, C. A.; Slavik, M.; Chien, S. C.;Hermann, J.; Thompson, G.; Blanc, O.; Invest. New Drugs (1987) 5, No. 2,177-86.

[0576] Phase II Trials of Alpha-Difluoromethylornithine, an Inhibitor ofPolyamine Synthesis, in Advanced Small Cell Lung Cancer and ColonCancer; Abeloff, M. D.; Rosen, S. T.; Luk, G. D.; Baylin, S. B.;Zeltzman, M.; Sjoerdsma, A. Cancer Treatment Reports, (1986) 70 (7)843-5.

[0577] Phase I Trial and Pharmacokinetic Studies of AlphaDifluoromethylomithine an Inhibitor of Poly Amine Biosynthesis; Abeloff,M. D.; Slavik, M.; Luk, G. D.; Griffin, C. A.; Hermann, J.; Blanc, O.;Sjoerdsma, A.; Baylin, S. B.; J. Clin. Oncol. (1984)2(2). pg. 124-130.

[0578] Regional Chemoprevention of Carcinogen-Induced Tumors in RatColon; Liu, T.; Mokuolu, A. O.; Rao, C. V.; Reddy, B. S.; Holt, P. R.;Gastroenterology, (1995) pg. 109/4 (1167-1172).

[0579] Placebo-Controlled Randomized Trial of DFMO As A ChemopreventiveAgent in Patients at High Risk For Colorectal Cancer; Jacoby, R. F.;Verma, A. K.; Tutsch, K. D.; Mamby, C. A.; Love, R. R.; Dept. ofMedicine, University of Wisconsin, Madison, Wis., U.S.A; J. Invest. Med.(1995) (43, Suppl. 2, 411A.)

[0580] Development of Difluoromethylornithine as a Chemoprevention Agentfor the Management of Colon Cancer. (Meyskens, F. L.; Gemer, E. W.;Department of Medicine and Clinical Cancer Center, University ofCalifornia, Irvine, 101 City Drive South, Building 23, Route 81, Orange,Calif. 92668, U.S.A. J. Cell. Biochem. (1995) (Suppl. 22, 126-31,) 3FIG. 2 Tab. 12 Ref.

[0581] Placebo-Controlled Randomized Trial of DFMO As A ChemopreventiveAgent in Patients at High Risk for Colorectal Cancer; Jacoby, R. F.;Verma, A. K.; Tutsch, K. D.; Mamby, C. A.; Love, R. R. Madison, Wis.,U.S.A.; Gastroenterology (108, No.4, Suppl., A485,)

[0582] Chronic Toxicity Studies of the Potential Cancer Preventive2-(difluoromethyl)-d,1-ornithine. (Crowell, J. A.; Goldenthal, E. I.;Kelloff, G. J.; Malone, W. F.; Boone, C. W. FUNDAM. APPL. TOXICOL.,(1994) 22i3 (341-354).

[0583] Oral Precancer: Preventive and Medical Approaches to Management;Scully, C., London, U. K.; Bur. J. Cancer Oral Oncol. (1995) (31, No. 1,pg. 16-26, 1

[0584] Chemopreventive Drug Development: Perspectives and Progress.Kelloff, G. J.; Boone, C. W.; Crowell, J. A.; Steele, V. E.; Lubet R.;Sigman C. C. Bethesda, M D. CIDU, National Cancer Institute, (1994)(85-98).

[0585] Chemoprevention of Barrett's Esophagus and Oral Leukoplakia.Garewal, H S; University of Arizona Cancer Center, Tucson. Adv Exp MedBiol, (1992) pg. 320 129-36.

[0586] Polyamines as Biomarkers of Cervical Intraepithelial NeoplasiaNishioka, Kenji; Melgarejo, Alejandro B.; Lyon, Rosanna R.; Mitchell,Michele Follen, Dep. of Surgical Oncology, Univ. of Texas, Houston,Tex., TX, 77030. J. Cell. Biochem. (1995), (Suppl. 23), (1995) pg. 87-95

[0587] Chemoprevention Trials and Surrogate End Point Biomarkers in theCervix; Mitchell, Michele Follen; Hittelman, Walter K.; Lotan, Reuben;Nishioka, Kenji; Tortolero-Luna, Guillermo; Richards-Kortum, Rebecca;Wharton, J. Taylor; Hong, Waun K. 1995Houston, Tex.) M. D. AndersonCancer Center, University Texas, Cancer (Philadelphia), (1995) pg.76(10, Suppl., American Cancer Society

[0588] Low Dose Difluoromethylornithine (DFMO) Produces SignificantChanges in Polyamine Content of Upper GI Mucosa in Patients withBarrett's Esophagus; Garewal, H S; Sampliner, R E; Fennerty, M B; Gemer,E., Tucson, Ariz.) Gastroenterology (100, No. 5, Pt. 2, A364), (1991) 1Ref.

[0589] Cancer Prevention Research Trials; Greenwald, P.; Malone, W. F.;Cerny, M. E.; Stem, H. R., Maryland 20892, U.S.A. 1993). Adv. CancerRes. (1993) (61, 1-23, 1993) 5 FIG. 4 Tab. 74 Ref.

[0590] Tumor Necrosis Factor-Induced Cytotoxicity is Accompanied byIntracellular Mitogenic Signals in ME-180Human Cervical Carcinoma Cells;Manchester, K. M.; Heston, W. D.; Donner, D. B.). Laboratory of PeptideHormone Action, Memorial, New York, N.Y. 10021.

[0591] Chemoprevention of Colon Carcinogenesis by Dietary Administrationof Piroxicam, .Alpha.-Difluoromethyl omithine,16.alpha.-Fluoro-5-androsten-17-one, and Ellagic Acid Individually andin Combination; Rao, Chinthalapally V.; Tokumo, Kenji; Rigotty, Jeff;Zang, Edith; Kelloff, Gary; Reddy, Bandaru S.; Div. Nutr. Carcinog.,American Health Found., Valhalla, N.Y., 10595, USA Cancer Res. (1991),51(17), 4528-34

[0592] Griffin C., Abeloff M D, Slavik M, et al. Phase I trial andpharmacokinetic study of intravenous and high dose oralα-difluoromethylomithine (DFMO). Proc ASCO 3:34, 1984.

[0593] Goldenthal, E. I. (1990). One Year Oral Toxicity Study ofDifluoromethylornithine in Rats and in Dogs. International Research andDevelopment Corporation, Reports 560-032 and 560-033.

[0594] Loprinzi, C. L., Love, R. R., Themeau, T. M., and Verma, A. K.;Inhibition of human skin ornithine decarboxylase activity by oraldifluoromethylornithine. Cancer Ther. Control 1, (1989) 75-80.

[0595] Luk, G. D.; Clinical and biologic studies of DFMO in the colon.In Cancer Chemoprevention (L. Wattenberg, M. Lipkin, C. W. Boone, and G.J. Kelloff, Eds.), (1992) pp. 515-530. CRC Press, Boca Raton, Fla.

[0596] Creaven P J, Pendyala L., Petreli N J: Evaluation of aα-difluoromethylornithine as a potential chemoprevention agent:Tolerance to daily oral administration in humans. Cancer EpidemiolBiomarkers Prev 2:243-247, (1993).

[0597] Croghan M K, Aickin M G, Meyakens Fla. Jr: Dose-relatedα-difluoromethylomithine ototoxicity. Am J Clin Oncol 14:331-335,(1991).

[0598] A Delayed Delivery System for the Colonic Drug Release; M. A.Vandelli et al., Proc. 1 st World Mtg. APGI/APV, Budapest, 9/11, May1995, pg. 278-279.

[0599] The Relation Between Swelling Properties and EnzymaticDegradation of Azo Polymers Designed for Colon-Specific Drug Delivery;G. Van den mooter et al., Pharmaceutical Res. (1994), 11 (12), pg.1737-1741.

[0600] Characterization of Colon-Specific Azopolymers: A Study of theSwelling Properties and the Permeability of Isolated Polymer Fiulms; G.Van den mooter et al., Internat'l. J. Pharmaceutics (1994), 111 pg.127-136.

[0601] Enteric Coated Timed Release Systems for Colonic Targeting; I. R.Wilding et al.; Internat'l. J. Pharmaceutics (1994), 111, pg. 99-102.

[0602] In Vivo Evaluation of a Colon-Specific Drug Delivery system: AnAbsorption Study of Theophylline from Capsules Coated with Azo Polymersin Rats; Pharmaceutical Res. (1995), 2(2), pg. 244-247.

[0603] Colonic Drug Delivery; T. N. Toer, Proceed. Intern. Symp. ControlRel. Bioact. Mater., March 16 (1990), pg. 126-127, pg. 291-295.

[0604] In Vitro and In Vivo Analysis of Colon Specificity of CalciumPectinate Formulations; A. Rubinstein et al., Eur. J. Pharm. Biopharm.(1995), 41(5), pg. 291-295.

What is claimed is:
 1. A pharmaceutical formulation comprising: a corehaving a rapid release DFMO-containing granules and a slow releasegranule, said granules comprising (+)-DFMO, (−)-DFMO or a defined ratiothereof, or pharmaceutically acceptable salts thereof; and an outerlayer surrounding said core comprising a pH responsive coating.
 2. Thepharmaceutical formulation of claim 1 wherein the rapid release granuleis capable of releasing DFMO within two hours after dissolution of theouter layer and the slow release granule is capable of releasing DFMOwithin eight hours after dissolution of the outer layer in an animal. 3.The pharmaceutical formulation of claim 1 or 2 wherein the pH responsivecoating is further defined as responsive to a pH of about 6.0.
 4. Anoral solid multiple drug release pharmaceutical formulation comprising:a core comprising DFMO-containing granules; and a pH-responsive outerlayer surrounding the core; wherein the granules comprise (+)-DFMO,(−)-DFMO or a defined ratio thereof, or pharmaceutically acceptablesalts thereof.
 5. The pharmaceutical formulation of claim 1 wherein theDFMO-containing granules are further defined as comprising: a gastricrelease granule; an enteric release granule; and a colorectal releasegranule, wherein each granule comprises (+)-DFMO, (−)-DFMO or a definedratio thereof, or pharmaceutically acceptable salts thereof.
 6. Apharmaceutical formulation core comprising: a rapid release granulehaving a binder, and a slow release granule having a polymer, eachgranule comprising (+)-DFMO, (−)-DFMO or a defined ratio thereof orpharmaceutically acceptable salts thereof; and an outer layersurrounding said granule comprising a pH-responsive coating, said pHresponsive coating providing for release of the core granules at orabove a pH of about
 6. 7. A multiple drug release profile pharmaceuticalformulation comprising: a core comprising a gastric release granulehaving a binder and an excipient; an enteric release granule comprisinga polymer suitable for enteric drug delivery; a colorectal releasegranule comprising a polymer which dissolves at a pH greater than orequal to about 6; and an outer layer surrounding the core, wherein eachgranule comprises a therapeutic amount of (+) DFMO, (−)-DFMOpharmaceutically acceptable salts thereof.
 8. The pharmaceuticalformulation of claim 1, 2 or 6 wherein the formulation is capable ofproviding a detectable plasma concentration of DFMO in the range ofabout 0.1 μM to about 1000 μM.
 9. The sustained release pharmaceuticalformulation of claim 8 wherein the formulation is capable of providing adetectable plasma concentration of DFMO in the range of about 1 μM toabout 100 μM.
 10. The solid multiple drug release profile pharmaceuticalformulation of claim 4 wherein the formulation is capable of providing aplasma concentration level of DFMO of about 0.1 μM to about 1000 μM. 11.The solid multiple drug release pharmaceutical formulation of claim 4further defined as an oral solid multiple drug release pharmaceuticalformulation, wherein the detectable plasma level of DFMO is about 1 μMto about 100 μM.
 12. A multi-coated pharmaceutical formulationcomprising a core pellet of DFMO-containing granules in a slow-releasematrix, a first coat of a colonic protective material; a second coatcomprising a pore forming agent; a rapidly dissolving polymeric materialand DFMO; a third coat comprising an acid resistant polymer; and afourth coat comprising a pore-forming agent and DFMO.
 13. Themulti-coated pharmaceutical formulation of claim 12 wherein theDFMO-containing granules comprise coated DFMO-containing granules anduncoated DFMO-containing granules.